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Search results for the GEO ID: GSE15999

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GSM400502

GPL1261

Pooled lung tissue (n=8) 6 hours after pneumonectomy

Lung tissue, 6 hours after left unilateral pneumonectomy

strain: C57BL/6 gender: Female age: 10-12 weeks treatment: left unilateral pneumonectomy time: 6 hours

Gene expression data from 8 pooled lung tissue samples 6 hours after left unilateral pneumonectomy

Sample_geo_accession	GSM400502
Sample_status	Public on May 08 2009
Sample_submission_date	May 06 2009
Sample_last_update_date	May 07 2009
Sample_type	RNA
Sample_channel_count	1
Sample_organism_ch1	Mus musculus
Sample_taxid_ch1	10090
Sample_treatment_protocol_ch1	Mice were anesthetized by intraperitoneal injection of ketamine (50-75 mg/kg) and xylazine (5 mg/kg), and then received 2ml of warmed normal saline and 100mg/kg sodium ampicillin subcutaneously. Orotracheal intubation was performed under direct visualization using a 20-gauge catheter (BD Insyte catheter; Becton, Dickinson and Co, Franklin Lakes NJ) over a flexible stylet. Mice were secured in supine position, and mechanically ventilated (AUT6110, Buxco Electronics, Wilmington, NC) at 200 tidal breaths of 0.3 ml of room air per minute, at positive end-expiratory pressure of 3 cm H2O during surgery and recovery. After achieving adequate anesthetic depth (absence of response to toe-pinch) the left thoracic wall was clipped and disinfected. The skin, chest wall and pleura were incised at the 5th intercostal space, and the left lung was gently lifted through a ~5-7mm incision and ligated at the hilum with 4-0 silk (Sofsilk, Synture Norwalk Ct). The lungs were then inflated to 30 cmH20 airway pressure, and the chest wall closed during this inflation with a single interrupted suture. The skin was closed with 5-0 PDS in a simple interrupted pattern. Mice were extubated at the onset of vigorous spontaneous breathing. The mice recovered from surgery in a warmed cage, and post-operative pain was managed with buprenorphine subcutaneously (0.05mg/kg) as soon as mice showed conscious motor control, and every 12 hours thereafter as needed (<3 days). Chow, nutrient gel (on the cage floor), and water were provided ad libitum. Sham pneumonectomy animals underwent an identical procedure, except that after the thoracotomy, the chest was left open for 5 minutes to simulate the conditions of the pneumonectomy group without removal of the left lung, then closed as described.
Sample_growth_protocol_ch1	Mice used in this study were adult (10-12 week) female C57BL/6 (20-25g) obtained from Jackson Laboratories.
Sample_molecule_ch1	total RNA
Sample_extract_protocol_ch1	The mice were anesthetized as above at 6 hours, 1 day, 3 days and 7 days after surgery (PNY or SHAM) then euthanized by cervical dislocation. The pulmonary vasculature was perfused with ice cold Hanks balanced salt solution, the trachea cannulated, and the lungs removed en bloc. RNA preservation was achieved by flooding the lung intratracheally with RNAlater solution (Qiagen #76104), followed by storage of lung tissue samples in RNAlater at -80ºC. Equal amounts of lung tissue were pooled from eight animals in each group (PNY or SHAM at each of 4 time points) to minimize biological variability. Total RNA was prepared from the dissected lung tissue using the Qiagen RNAeasy mini kit (Qiagen #74104) according to the manufacturer’s directions. The total RNA samples from the primary purification were purified a second time on Qiagen RNAeasy columns according to the manufacturer’s instructions. Total RNA concentrations, A260/A280 and A260/A230 ratios was determined using a NanoDrop ND1000 spectrophotometer.
Sample_label_ch1	Biotin
Sample_label_protocol_ch1	Labeling was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_hyb_protocol	Chip hybridization was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_scan_protocol	GeneChips were scanned using the Affymetrix 7G scanner.
Sample_data_processing = The initial goal in the analysis was to identify genes regulated in the comparison between pneumonectomized and sham-operated animals. By treating all time points as replicates within their respective groups two datasets were created (PNY and SHAM), each with an n	4. The corresponding Affymetrix CEL files were background corrected, summarized and quantile-normalized using the RMA library in BioConductor (http:www.bioconductor.org), yielding one expression value per probe set for each of the 8 arrays. Based on the ‘Rank Products’ algorithm proposed by Brietling, et. al, the RankProd library was employed to find differentially expressed genes. This algorithm works by performing comprehensive pair-wise comparisons to calculate a rank statistic RPg, the probability of seeing the observed, pair-wise expression patterns for any given gene g. As a vehicle for measuring statistical significance a non-parametric P-value is also calculated, using 1000 permutations to determine how often the calculated RPg statistic would occur by chance alone. Finally, the RankProd library compares average expression between the two groups to derive a fold-change value. Genes with a reported P-value < 0.001 and a fold-change > 1.5 or < -1.5 were selected for further investigation. In a second analysis, the focus was shifted to temporal changes in gene expression during lung regeneration as opposed to overall transcriptomic patterns. With only one array per experimental condition at each time point, derivation of statistical measures and the subsequent search for truly differentially expressed genes can be challenging. However, the S-Score algorithm described by Zhang et al. (2002) and Kerns et al. (2003) provides a method for determining statistical significance when biological replicates are not available by applying pairwise comparisons to probe-level data. On average, the Affymetrix 3’ IVT platform contains 22 probes for every transcript represented on the array. Using this information directly, the S-Score algorithm has shown an increased sensitivity when compared to many other existing analysis methods without sacrificing specificity, and can produce accurate results when no biological replicates are present (This is particularly applicable and appropriate to our individual time point datasets in which we have only one paired array set for each time point. Using the S-Score algorithm, the relative change in probe pair intensity is calculated to convert the probe pair signal differences into multiple measurements with equalized errors. The relative changes for each probe pair are then summed to form the S-score, which represents a single measure of the significance of change for the gene in question. By definition, S-score is related to P-value by an exponential relation, and a value of 3 corresponds to a P-value of 0.003 . Therefore, genes with an S-score > 3.0 or < - 3.0 (P < 0.003) were selected for further analysis.
Sample_platform_id	GPL1261
Sample_contact_name	Julia,A,Paxson
Sample_contact_email	julia.paxson@tufts.edu
Sample_contact_laboratory	Hoffman Lab
Sample_contact_department	Clinical Sciences
Sample_contact_institute	Tufts Cummings School of Veterinary Medicine
Sample_contact_address	200 Westboro Road
Sample_contact_city	North Grafton
Sample_contact_state	MA
Sample_contact_zip/postal_code	01536
Sample_contact_country	USA
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM400nnn/GSM400502/suppl/GSM400502.CEL.gz
Sample_series_id	GSE15999
Sample_data_row_count	45101

GSM400503

GPL1261

Pooled lung tissue (n=8) 6 hours after sham thoracotomy

Lung tissue, 6 hours after sham thoracotomy

strain: C57BL/6 gender: Female age: 10-12 weeks treatment: sham thoracotomy time: 6 hours

Gene expression data from 8 pooled lung tissue samples 6 hours after sham thoracotomy

Sample_geo_accession	GSM400503
Sample_status	Public on May 08 2009
Sample_submission_date	May 06 2009
Sample_last_update_date	May 07 2009
Sample_type	RNA
Sample_channel_count	1
Sample_organism_ch1	Mus musculus
Sample_taxid_ch1	10090
Sample_treatment_protocol_ch1	Mice were anesthetized by intraperitoneal injection of ketamine (50-75 mg/kg) and xylazine (5 mg/kg), and then received 2ml of warmed normal saline and 100mg/kg sodium ampicillin subcutaneously. Orotracheal intubation was performed under direct visualization using a 20-gauge catheter (BD Insyte catheter; Becton, Dickinson and Co, Franklin Lakes NJ) over a flexible stylet. Mice were secured in supine position, and mechanically ventilated (AUT6110, Buxco Electronics, Wilmington, NC) at 200 tidal breaths of 0.3 ml of room air per minute, at positive end-expiratory pressure of 3 cm H2O during surgery and recovery. After achieving adequate anesthetic depth (absence of response to toe-pinch) the left thoracic wall was clipped and disinfected. The skin, chest wall and pleura were incised at the 5th intercostal space, and the left lung was gently lifted through a ~5-7mm incision and ligated at the hilum with 4-0 silk (Sofsilk, Synture Norwalk Ct). The lungs were then inflated to 30 cmH20 airway pressure, and the chest wall closed during this inflation with a single interrupted suture. The skin was closed with 5-0 PDS in a simple interrupted pattern. Mice were extubated at the onset of vigorous spontaneous breathing. The mice recovered from surgery in a warmed cage, and post-operative pain was managed with buprenorphine subcutaneously (0.05mg/kg) as soon as mice showed conscious motor control, and every 12 hours thereafter as needed (<3 days). Chow, nutrient gel (on the cage floor), and water were provided ad libitum. Sham pneumonectomy animals underwent an identical procedure, except that after the thoracotomy, the chest was left open for 5 minutes to simulate the conditions of the pneumonectomy group without removal of the left lung, then closed as described.
Sample_growth_protocol_ch1	Mice used in this study were adult (10-12 week) female C57BL/6 (20-25g) obtained from Jackson Laboratories.
Sample_molecule_ch1	total RNA
Sample_extract_protocol_ch1	The mice were anesthetized as above at 6 hours, 1 day, 3 days and 7 days after surgery (PNY or SHAM) then euthanized by cervical dislocation. The pulmonary vasculature was perfused with ice cold Hanks balanced salt solution, the trachea cannulated, and the lungs removed en bloc. RNA preservation was achieved by flooding the lung intratracheally with RNAlater solution (Qiagen #76104), followed by storage of lung tissue samples in RNAlater at -80ºC. Equal amounts of lung tissue were pooled from eight animals in each group (PNY or SHAM at each of 4 time points) to minimize biological variability. Total RNA was prepared from the dissected lung tissue using the Qiagen RNAeasy mini kit (Qiagen #74104) according to the manufacturer’s directions. The total RNA samples from the primary purification were purified a second time on Qiagen RNAeasy columns according to the manufacturer’s instructions. Total RNA concentrations, A260/A280 and A260/A230 ratios was determined using a NanoDrop ND1000 spectrophotometer.
Sample_label_ch1	Biotin
Sample_label_protocol_ch1	Labeling was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_hyb_protocol	Chip hybridization was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_scan_protocol	GeneChips were scanned using the Affymetrix 7G scanner.
Sample_data_processing = The initial goal in the analysis was to identify genes regulated in the comparison between pneumonectomized and sham-operated animals. By treating all time points as replicates within their respective groups two datasets were created (PNY and SHAM), each with an n	4. The corresponding Affymetrix CEL files were background corrected, summarized and quantile-normalized using the RMA library in BioConductor (http:www.bioconductor.org), yielding one expression value per probe set for each of the 8 arrays. Based on the ‘Rank Products’ algorithm proposed by Brietling, et. al, the RankProd library was employed to find differentially expressed genes. This algorithm works by performing comprehensive pair-wise comparisons to calculate a rank statistic RPg, the probability of seeing the observed, pair-wise expression patterns for any given gene g. As a vehicle for measuring statistical significance a non-parametric P-value is also calculated, using 1000 permutations to determine how often the calculated RPg statistic would occur by chance alone. Finally, the RankProd library compares average expression between the two groups to derive a fold-change value. Genes with a reported P-value < 0.001 and a fold-change > 1.5 or < -1.5 were selected for further investigation. In a second analysis, the focus was shifted to temporal changes in gene expression during lung regeneration as opposed to overall transcriptomic patterns. With only one array per experimental condition at each time point, derivation of statistical measures and the subsequent search for truly differentially expressed genes can be challenging. However, the S-Score algorithm described by Zhang et al. (2002) and Kerns et al. (2003) provides a method for determining statistical significance when biological replicates are not available by applying pairwise comparisons to probe-level data. On average, the Affymetrix 3’ IVT platform contains 22 probes for every transcript represented on the array. Using this information directly, the S-Score algorithm has shown an increased sensitivity when compared to many other existing analysis methods without sacrificing specificity, and can produce accurate results when no biological replicates are present (This is particularly applicable and appropriate to our individual time point datasets in which we have only one paired array set for each time point. Using the S-Score algorithm, the relative change in probe pair intensity is calculated to convert the probe pair signal differences into multiple measurements with equalized errors. The relative changes for each probe pair are then summed to form the S-score, which represents a single measure of the significance of change for the gene in question. By definition, S-score is related to P-value by an exponential relation, and a value of 3 corresponds to a P-value of 0.003 . Therefore, genes with an S-score > 3.0 or < - 3.0 (P < 0.003) were selected for further analysis.
Sample_platform_id	GPL1261
Sample_contact_name	Julia,A,Paxson
Sample_contact_email	julia.paxson@tufts.edu
Sample_contact_laboratory	Hoffman Lab
Sample_contact_department	Clinical Sciences
Sample_contact_institute	Tufts Cummings School of Veterinary Medicine
Sample_contact_address	200 Westboro Road
Sample_contact_city	North Grafton
Sample_contact_state	MA
Sample_contact_zip/postal_code	01536
Sample_contact_country	USA
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM400nnn/GSM400503/suppl/GSM400503.CEL.gz
Sample_series_id	GSE15999
Sample_data_row_count	45101

GSM400504

GPL1261

Pooled lung tissue (n=8) 1 day after pneumonectomy

Lung tissue, 1 day after left unilateral pneumonectomy

strain: C57BL/6 gender: Female age: 10-12 weeks treatment: left unilateral pneumonectomy time: 1 day

Gene expression data from 8 pooled lung tissue samples 1 day after left unilateral pneumonectomy

Sample_geo_accession	GSM400504
Sample_status	Public on May 08 2009
Sample_submission_date	May 06 2009
Sample_last_update_date	May 07 2009
Sample_type	RNA
Sample_channel_count	1
Sample_organism_ch1	Mus musculus
Sample_taxid_ch1	10090
Sample_treatment_protocol_ch1	Mice were anesthetized by intraperitoneal injection of ketamine (50-75 mg/kg) and xylazine (5 mg/kg), and then received 2ml of warmed normal saline and 100mg/kg sodium ampicillin subcutaneously. Orotracheal intubation was performed under direct visualization using a 20-gauge catheter (BD Insyte catheter; Becton, Dickinson and Co, Franklin Lakes NJ) over a flexible stylet. Mice were secured in supine position, and mechanically ventilated (AUT6110, Buxco Electronics, Wilmington, NC) at 200 tidal breaths of 0.3 ml of room air per minute, at positive end-expiratory pressure of 3 cm H2O during surgery and recovery. After achieving adequate anesthetic depth (absence of response to toe-pinch) the left thoracic wall was clipped and disinfected. The skin, chest wall and pleura were incised at the 5th intercostal space, and the left lung was gently lifted through a ~5-7mm incision and ligated at the hilum with 4-0 silk (Sofsilk, Synture Norwalk Ct). The lungs were then inflated to 30 cmH20 airway pressure, and the chest wall closed during this inflation with a single interrupted suture. The skin was closed with 5-0 PDS in a simple interrupted pattern. Mice were extubated at the onset of vigorous spontaneous breathing. The mice recovered from surgery in a warmed cage, and post-operative pain was managed with buprenorphine subcutaneously (0.05mg/kg) as soon as mice showed conscious motor control, and every 12 hours thereafter as needed (<3 days). Chow, nutrient gel (on the cage floor), and water were provided ad libitum. Sham pneumonectomy animals underwent an identical procedure, except that after the thoracotomy, the chest was left open for 5 minutes to simulate the conditions of the pneumonectomy group without removal of the left lung, then closed as described.
Sample_growth_protocol_ch1	Mice used in this study were adult (10-12 week) female C57BL/6 (20-25g) obtained from Jackson Laboratories.
Sample_molecule_ch1	total RNA
Sample_extract_protocol_ch1	The mice were anesthetized as above at 6 hours, 1 day, 3 days and 7 days after surgery (PNY or SHAM) then euthanized by cervical dislocation. The pulmonary vasculature was perfused with ice cold Hanks balanced salt solution, the trachea cannulated, and the lungs removed en bloc. RNA preservation was achieved by flooding the lung intratracheally with RNAlater solution (Qiagen #76104), followed by storage of lung tissue samples in RNAlater at -80ºC. Equal amounts of lung tissue were pooled from eight animals in each group (PNY or SHAM at each of 4 time points) to minimize biological variability. Total RNA was prepared from the dissected lung tissue using the Qiagen RNAeasy mini kit (Qiagen #74104) according to the manufacturer’s directions. The total RNA samples from the primary purification were purified a second time on Qiagen RNAeasy columns according to the manufacturer’s instructions. Total RNA concentrations, A260/A280 and A260/A230 ratios was determined using a NanoDrop ND1000 spectrophotometer.
Sample_label_ch1	Biotin
Sample_label_protocol_ch1	Labeling was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_hyb_protocol	Chip hybridization was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_scan_protocol	GeneChips were scanned using the Affymetrix 7G scanner.
Sample_data_processing = The initial goal in the analysis was to identify genes regulated in the comparison between pneumonectomized and sham-operated animals. By treating all time points as replicates within their respective groups two datasets were created (PNY and SHAM), each with an n	4. The corresponding Affymetrix CEL files were background corrected, summarized and quantile-normalized using the RMA library in BioConductor (http:www.bioconductor.org), yielding one expression value per probe set for each of the 8 arrays. Based on the ‘Rank Products’ algorithm proposed by Brietling, et. al, the RankProd library was employed to find differentially expressed genes. This algorithm works by performing comprehensive pair-wise comparisons to calculate a rank statistic RPg, the probability of seeing the observed, pair-wise expression patterns for any given gene g. As a vehicle for measuring statistical significance a non-parametric P-value is also calculated, using 1000 permutations to determine how often the calculated RPg statistic would occur by chance alone. Finally, the RankProd library compares average expression between the two groups to derive a fold-change value. Genes with a reported P-value < 0.001 and a fold-change > 1.5 or < -1.5 were selected for further investigation. In a second analysis, the focus was shifted to temporal changes in gene expression during lung regeneration as opposed to overall transcriptomic patterns. With only one array per experimental condition at each time point, derivation of statistical measures and the subsequent search for truly differentially expressed genes can be challenging. However, the S-Score algorithm described by Zhang et al. (2002) and Kerns et al. (2003) provides a method for determining statistical significance when biological replicates are not available by applying pairwise comparisons to probe-level data. On average, the Affymetrix 3’ IVT platform contains 22 probes for every transcript represented on the array. Using this information directly, the S-Score algorithm has shown an increased sensitivity when compared to many other existing analysis methods without sacrificing specificity, and can produce accurate results when no biological replicates are present (This is particularly applicable and appropriate to our individual time point datasets in which we have only one paired array set for each time point. Using the S-Score algorithm, the relative change in probe pair intensity is calculated to convert the probe pair signal differences into multiple measurements with equalized errors. The relative changes for each probe pair are then summed to form the S-score, which represents a single measure of the significance of change for the gene in question. By definition, S-score is related to P-value by an exponential relation, and a value of 3 corresponds to a P-value of 0.003 . Therefore, genes with an S-score > 3.0 or < - 3.0 (P < 0.003) were selected for further analysis.
Sample_platform_id	GPL1261
Sample_contact_name	Julia,A,Paxson
Sample_contact_email	julia.paxson@tufts.edu
Sample_contact_laboratory	Hoffman Lab
Sample_contact_department	Clinical Sciences
Sample_contact_institute	Tufts Cummings School of Veterinary Medicine
Sample_contact_address	200 Westboro Road
Sample_contact_city	North Grafton
Sample_contact_state	MA
Sample_contact_zip/postal_code	01536
Sample_contact_country	USA
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM400nnn/GSM400504/suppl/GSM400504.CEL.gz
Sample_series_id	GSE15999
Sample_data_row_count	45101

GSM400505

GPL1261

Pooled lung tissue (n=8) 1 day after sham thoracotomy

Lung tissue, 1 day after sham thoracotomy

strain: C57BL/6 gender: Female age: 10-12 weeks treatment: sham thoracotomy time: 1 day

Gene expression data from 8 pooled lung tissue samples 1 day after sham thoracotomy

Sample_geo_accession	GSM400505
Sample_status	Public on May 08 2009
Sample_submission_date	May 06 2009
Sample_last_update_date	May 07 2009
Sample_type	RNA
Sample_channel_count	1
Sample_organism_ch1	Mus musculus
Sample_taxid_ch1	10090
Sample_treatment_protocol_ch1	Mice were anesthetized by intraperitoneal injection of ketamine (50-75 mg/kg) and xylazine (5 mg/kg), and then received 2ml of warmed normal saline and 100mg/kg sodium ampicillin subcutaneously. Orotracheal intubation was performed under direct visualization using a 20-gauge catheter (BD Insyte catheter; Becton, Dickinson and Co, Franklin Lakes NJ) over a flexible stylet. Mice were secured in supine position, and mechanically ventilated (AUT6110, Buxco Electronics, Wilmington, NC) at 200 tidal breaths of 0.3 ml of room air per minute, at positive end-expiratory pressure of 3 cm H2O during surgery and recovery. After achieving adequate anesthetic depth (absence of response to toe-pinch) the left thoracic wall was clipped and disinfected. The skin, chest wall and pleura were incised at the 5th intercostal space, and the left lung was gently lifted through a ~5-7mm incision and ligated at the hilum with 4-0 silk (Sofsilk, Synture Norwalk Ct). The lungs were then inflated to 30 cmH20 airway pressure, and the chest wall closed during this inflation with a single interrupted suture. The skin was closed with 5-0 PDS in a simple interrupted pattern. Mice were extubated at the onset of vigorous spontaneous breathing. The mice recovered from surgery in a warmed cage, and post-operative pain was managed with buprenorphine subcutaneously (0.05mg/kg) as soon as mice showed conscious motor control, and every 12 hours thereafter as needed (<3 days). Chow, nutrient gel (on the cage floor), and water were provided ad libitum. Sham pneumonectomy animals underwent an identical procedure, except that after the thoracotomy, the chest was left open for 5 minutes to simulate the conditions of the pneumonectomy group without removal of the left lung, then closed as described.
Sample_growth_protocol_ch1	Mice used in this study were adult (10-12 week) female C57BL/6 (20-25g) obtained from Jackson Laboratories.
Sample_molecule_ch1	total RNA
Sample_extract_protocol_ch1	The mice were anesthetized as above at 6 hours, 1 day, 3 days and 7 days after surgery (PNY or SHAM) then euthanized by cervical dislocation. The pulmonary vasculature was perfused with ice cold Hanks balanced salt solution, the trachea cannulated, and the lungs removed en bloc. RNA preservation was achieved by flooding the lung intratracheally with RNAlater solution (Qiagen #76104), followed by storage of lung tissue samples in RNAlater at -80ºC. Equal amounts of lung tissue were pooled from eight animals in each group (PNY or SHAM at each of 4 time points) to minimize biological variability. Total RNA was prepared from the dissected lung tissue using the Qiagen RNAeasy mini kit (Qiagen #74104) according to the manufacturer’s directions. The total RNA samples from the primary purification were purified a second time on Qiagen RNAeasy columns according to the manufacturer’s instructions. Total RNA concentrations, A260/A280 and A260/A230 ratios was determined using a NanoDrop ND1000 spectrophotometer.
Sample_label_ch1	Biotin
Sample_label_protocol_ch1	Labeling was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_hyb_protocol	Chip hybridization was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_scan_protocol	GeneChips were scanned using the Affymetrix 7G scanner.
Sample_data_processing = The initial goal in the analysis was to identify genes regulated in the comparison between pneumonectomized and sham-operated animals. By treating all time points as replicates within their respective groups two datasets were created (PNY and SHAM), each with an n	4. The corresponding Affymetrix CEL files were background corrected, summarized and quantile-normalized using the RMA library in BioConductor (http:www.bioconductor.org), yielding one expression value per probe set for each of the 8 arrays. Based on the ‘Rank Products’ algorithm proposed by Brietling, et. al, the RankProd library was employed to find differentially expressed genes. This algorithm works by performing comprehensive pair-wise comparisons to calculate a rank statistic RPg, the probability of seeing the observed, pair-wise expression patterns for any given gene g. As a vehicle for measuring statistical significance a non-parametric P-value is also calculated, using 1000 permutations to determine how often the calculated RPg statistic would occur by chance alone. Finally, the RankProd library compares average expression between the two groups to derive a fold-change value. Genes with a reported P-value < 0.001 and a fold-change > 1.5 or < -1.5 were selected for further investigation. In a second analysis, the focus was shifted to temporal changes in gene expression during lung regeneration as opposed to overall transcriptomic patterns. With only one array per experimental condition at each time point, derivation of statistical measures and the subsequent search for truly differentially expressed genes can be challenging. However, the S-Score algorithm described by Zhang et al. (2002) and Kerns et al. (2003) provides a method for determining statistical significance when biological replicates are not available by applying pairwise comparisons to probe-level data. On average, the Affymetrix 3’ IVT platform contains 22 probes for every transcript represented on the array. Using this information directly, the S-Score algorithm has shown an increased sensitivity when compared to many other existing analysis methods without sacrificing specificity, and can produce accurate results when no biological replicates are present (This is particularly applicable and appropriate to our individual time point datasets in which we have only one paired array set for each time point. Using the S-Score algorithm, the relative change in probe pair intensity is calculated to convert the probe pair signal differences into multiple measurements with equalized errors. The relative changes for each probe pair are then summed to form the S-score, which represents a single measure of the significance of change for the gene in question. By definition, S-score is related to P-value by an exponential relation, and a value of 3 corresponds to a P-value of 0.003 . Therefore, genes with an S-score > 3.0 or < - 3.0 (P < 0.003) were selected for further analysis.
Sample_platform_id	GPL1261
Sample_contact_name	Julia,A,Paxson
Sample_contact_email	julia.paxson@tufts.edu
Sample_contact_laboratory	Hoffman Lab
Sample_contact_department	Clinical Sciences
Sample_contact_institute	Tufts Cummings School of Veterinary Medicine
Sample_contact_address	200 Westboro Road
Sample_contact_city	North Grafton
Sample_contact_state	MA
Sample_contact_zip/postal_code	01536
Sample_contact_country	USA
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM400nnn/GSM400505/suppl/GSM400505.CEL.gz
Sample_series_id	GSE15999
Sample_data_row_count	45101

GSM400506

GPL1261

Pooled lung tissue (n=8) 3 days after pneumonectomy

Lung tissue, 3 days after left unilateral pneumonectomy

strain: C57BL/6 gender: Female age: 10-12 weeks treatment: left unilateral pneumonectomy time: 3 days

Gene expression data from 8 pooled lung tissue samples 3 days after left unilateral pneumonectomy

Sample_geo_accession	GSM400506
Sample_status	Public on May 08 2009
Sample_submission_date	May 06 2009
Sample_last_update_date	May 07 2009
Sample_type	RNA
Sample_channel_count	1
Sample_organism_ch1	Mus musculus
Sample_taxid_ch1	10090
Sample_treatment_protocol_ch1	Mice were anesthetized by intraperitoneal injection of ketamine (50-75 mg/kg) and xylazine (5 mg/kg), and then received 2ml of warmed normal saline and 100mg/kg sodium ampicillin subcutaneously. Orotracheal intubation was performed under direct visualization using a 20-gauge catheter (BD Insyte catheter; Becton, Dickinson and Co, Franklin Lakes NJ) over a flexible stylet. Mice were secured in supine position, and mechanically ventilated (AUT6110, Buxco Electronics, Wilmington, NC) at 200 tidal breaths of 0.3 ml of room air per minute, at positive end-expiratory pressure of 3 cm H2O during surgery and recovery. After achieving adequate anesthetic depth (absence of response to toe-pinch) the left thoracic wall was clipped and disinfected. The skin, chest wall and pleura were incised at the 5th intercostal space, and the left lung was gently lifted through a ~5-7mm incision and ligated at the hilum with 4-0 silk (Sofsilk, Synture Norwalk Ct). The lungs were then inflated to 30 cmH20 airway pressure, and the chest wall closed during this inflation with a single interrupted suture. The skin was closed with 5-0 PDS in a simple interrupted pattern. Mice were extubated at the onset of vigorous spontaneous breathing. The mice recovered from surgery in a warmed cage, and post-operative pain was managed with buprenorphine subcutaneously (0.05mg/kg) as soon as mice showed conscious motor control, and every 12 hours thereafter as needed (<3 days). Chow, nutrient gel (on the cage floor), and water were provided ad libitum. Sham pneumonectomy animals underwent an identical procedure, except that after the thoracotomy, the chest was left open for 5 minutes to simulate the conditions of the pneumonectomy group without removal of the left lung, then closed as described.
Sample_growth_protocol_ch1	Mice used in this study were adult (10-12 week) female C57BL/6 (20-25g) obtained from Jackson Laboratories.
Sample_molecule_ch1	total RNA
Sample_extract_protocol_ch1	The mice were anesthetized as above at 6 hours, 1 day, 3 days and 7 days after surgery (PNY or SHAM) then euthanized by cervical dislocation. The pulmonary vasculature was perfused with ice cold Hanks balanced salt solution, the trachea cannulated, and the lungs removed en bloc. RNA preservation was achieved by flooding the lung intratracheally with RNAlater solution (Qiagen #76104), followed by storage of lung tissue samples in RNAlater at -80ºC. Equal amounts of lung tissue were pooled from eight animals in each group (PNY or SHAM at each of 4 time points) to minimize biological variability. Total RNA was prepared from the dissected lung tissue using the Qiagen RNAeasy mini kit (Qiagen #74104) according to the manufacturer’s directions. The total RNA samples from the primary purification were purified a second time on Qiagen RNAeasy columns according to the manufacturer’s instructions. Total RNA concentrations, A260/A280 and A260/A230 ratios was determined using a NanoDrop ND1000 spectrophotometer.
Sample_label_ch1	Biotin
Sample_label_protocol_ch1	Labeling was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_hyb_protocol	Chip hybridization was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_scan_protocol	GeneChips were scanned using the Affymetrix 7G scanner.
Sample_data_processing = The initial goal in the analysis was to identify genes regulated in the comparison between pneumonectomized and sham-operated animals. By treating all time points as replicates within their respective groups two datasets were created (PNY and SHAM), each with an n	4. The corresponding Affymetrix CEL files were background corrected, summarized and quantile-normalized using the RMA library in BioConductor (http:www.bioconductor.org), yielding one expression value per probe set for each of the 8 arrays. Based on the ‘Rank Products’ algorithm proposed by Brietling, et. al, the RankProd library was employed to find differentially expressed genes. This algorithm works by performing comprehensive pair-wise comparisons to calculate a rank statistic RPg, the probability of seeing the observed, pair-wise expression patterns for any given gene g. As a vehicle for measuring statistical significance a non-parametric P-value is also calculated, using 1000 permutations to determine how often the calculated RPg statistic would occur by chance alone. Finally, the RankProd library compares average expression between the two groups to derive a fold-change value. Genes with a reported P-value < 0.001 and a fold-change > 1.5 or < -1.5 were selected for further investigation. In a second analysis, the focus was shifted to temporal changes in gene expression during lung regeneration as opposed to overall transcriptomic patterns. With only one array per experimental condition at each time point, derivation of statistical measures and the subsequent search for truly differentially expressed genes can be challenging. However, the S-Score algorithm described by Zhang et al. (2002) and Kerns et al. (2003) provides a method for determining statistical significance when biological replicates are not available by applying pairwise comparisons to probe-level data. On average, the Affymetrix 3’ IVT platform contains 22 probes for every transcript represented on the array. Using this information directly, the S-Score algorithm has shown an increased sensitivity when compared to many other existing analysis methods without sacrificing specificity, and can produce accurate results when no biological replicates are present (This is particularly applicable and appropriate to our individual time point datasets in which we have only one paired array set for each time point. Using the S-Score algorithm, the relative change in probe pair intensity is calculated to convert the probe pair signal differences into multiple measurements with equalized errors. The relative changes for each probe pair are then summed to form the S-score, which represents a single measure of the significance of change for the gene in question. By definition, S-score is related to P-value by an exponential relation, and a value of 3 corresponds to a P-value of 0.003 . Therefore, genes with an S-score > 3.0 or < - 3.0 (P < 0.003) were selected for further analysis.
Sample_platform_id	GPL1261
Sample_contact_name	Julia,A,Paxson
Sample_contact_email	julia.paxson@tufts.edu
Sample_contact_laboratory	Hoffman Lab
Sample_contact_department	Clinical Sciences
Sample_contact_institute	Tufts Cummings School of Veterinary Medicine
Sample_contact_address	200 Westboro Road
Sample_contact_city	North Grafton
Sample_contact_state	MA
Sample_contact_zip/postal_code	01536
Sample_contact_country	USA
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM400nnn/GSM400506/suppl/GSM400506.CEL.gz
Sample_series_id	GSE15999
Sample_data_row_count	45101

GSM400507

GPL1261

Pooled lung tissue (n=8) 3 days after sham thoracotomy

Lung tissue, 3 days after sham thoracotomy

strain: C57BL/6 gender: Female age: 10-12 weeks treatment: sham thoracotomy time: 3 days

Gene expression data from 8 pooled lung tissue samples 3 days after sham thoracotomy

Sample_geo_accession	GSM400507
Sample_status	Public on May 08 2009
Sample_submission_date	May 06 2009
Sample_last_update_date	May 07 2009
Sample_type	RNA
Sample_channel_count	1
Sample_organism_ch1	Mus musculus
Sample_taxid_ch1	10090
Sample_treatment_protocol_ch1	Mice were anesthetized by intraperitoneal injection of ketamine (50-75 mg/kg) and xylazine (5 mg/kg), and then received 2ml of warmed normal saline and 100mg/kg sodium ampicillin subcutaneously. Orotracheal intubation was performed under direct visualization using a 20-gauge catheter (BD Insyte catheter; Becton, Dickinson and Co, Franklin Lakes NJ) over a flexible stylet. Mice were secured in supine position, and mechanically ventilated (AUT6110, Buxco Electronics, Wilmington, NC) at 200 tidal breaths of 0.3 ml of room air per minute, at positive end-expiratory pressure of 3 cm H2O during surgery and recovery. After achieving adequate anesthetic depth (absence of response to toe-pinch) the left thoracic wall was clipped and disinfected. The skin, chest wall and pleura were incised at the 5th intercostal space, and the left lung was gently lifted through a ~5-7mm incision and ligated at the hilum with 4-0 silk (Sofsilk, Synture Norwalk Ct). The lungs were then inflated to 30 cmH20 airway pressure, and the chest wall closed during this inflation with a single interrupted suture. The skin was closed with 5-0 PDS in a simple interrupted pattern. Mice were extubated at the onset of vigorous spontaneous breathing. The mice recovered from surgery in a warmed cage, and post-operative pain was managed with buprenorphine subcutaneously (0.05mg/kg) as soon as mice showed conscious motor control, and every 12 hours thereafter as needed (<3 days). Chow, nutrient gel (on the cage floor), and water were provided ad libitum. Sham pneumonectomy animals underwent an identical procedure, except that after the thoracotomy, the chest was left open for 5 minutes to simulate the conditions of the pneumonectomy group without removal of the left lung, then closed as described.
Sample_growth_protocol_ch1	Mice used in this study were adult (10-12 week) female C57BL/6 (20-25g) obtained from Jackson Laboratories.
Sample_molecule_ch1	total RNA
Sample_extract_protocol_ch1	The mice were anesthetized as above at 6 hours, 1 day, 3 days and 7 days after surgery (PNY or SHAM) then euthanized by cervical dislocation. The pulmonary vasculature was perfused with ice cold Hanks balanced salt solution, the trachea cannulated, and the lungs removed en bloc. RNA preservation was achieved by flooding the lung intratracheally with RNAlater solution (Qiagen #76104), followed by storage of lung tissue samples in RNAlater at -80ºC. Equal amounts of lung tissue were pooled from eight animals in each group (PNY or SHAM at each of 4 time points) to minimize biological variability. Total RNA was prepared from the dissected lung tissue using the Qiagen RNAeasy mini kit (Qiagen #74104) according to the manufacturer’s directions. The total RNA samples from the primary purification were purified a second time on Qiagen RNAeasy columns according to the manufacturer’s instructions. Total RNA concentrations, A260/A280 and A260/A230 ratios was determined using a NanoDrop ND1000 spectrophotometer.
Sample_label_ch1	Biotin
Sample_label_protocol_ch1	Labeling was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_hyb_protocol	Chip hybridization was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_scan_protocol	GeneChips were scanned using the Affymetrix 7G scanner.
Sample_data_processing = The initial goal in the analysis was to identify genes regulated in the comparison between pneumonectomized and sham-operated animals. By treating all time points as replicates within their respective groups two datasets were created (PNY and SHAM), each with an n	4. The corresponding Affymetrix CEL files were background corrected, summarized and quantile-normalized using the RMA library in BioConductor (http:www.bioconductor.org), yielding one expression value per probe set for each of the 8 arrays. Based on the ‘Rank Products’ algorithm proposed by Brietling, et. al, the RankProd library was employed to find differentially expressed genes. This algorithm works by performing comprehensive pair-wise comparisons to calculate a rank statistic RPg, the probability of seeing the observed, pair-wise expression patterns for any given gene g. As a vehicle for measuring statistical significance a non-parametric P-value is also calculated, using 1000 permutations to determine how often the calculated RPg statistic would occur by chance alone. Finally, the RankProd library compares average expression between the two groups to derive a fold-change value. Genes with a reported P-value < 0.001 and a fold-change > 1.5 or < -1.5 were selected for further investigation. In a second analysis, the focus was shifted to temporal changes in gene expression during lung regeneration as opposed to overall transcriptomic patterns. With only one array per experimental condition at each time point, derivation of statistical measures and the subsequent search for truly differentially expressed genes can be challenging. However, the S-Score algorithm described by Zhang et al. (2002) and Kerns et al. (2003) provides a method for determining statistical significance when biological replicates are not available by applying pairwise comparisons to probe-level data. On average, the Affymetrix 3’ IVT platform contains 22 probes for every transcript represented on the array. Using this information directly, the S-Score algorithm has shown an increased sensitivity when compared to many other existing analysis methods without sacrificing specificity, and can produce accurate results when no biological replicates are present (This is particularly applicable and appropriate to our individual time point datasets in which we have only one paired array set for each time point. Using the S-Score algorithm, the relative change in probe pair intensity is calculated to convert the probe pair signal differences into multiple measurements with equalized errors. The relative changes for each probe pair are then summed to form the S-score, which represents a single measure of the significance of change for the gene in question. By definition, S-score is related to P-value by an exponential relation, and a value of 3 corresponds to a P-value of 0.003 . Therefore, genes with an S-score > 3.0 or < - 3.0 (P < 0.003) were selected for further analysis.
Sample_platform_id	GPL1261
Sample_contact_name	Julia,A,Paxson
Sample_contact_email	julia.paxson@tufts.edu
Sample_contact_laboratory	Hoffman Lab
Sample_contact_department	Clinical Sciences
Sample_contact_institute	Tufts Cummings School of Veterinary Medicine
Sample_contact_address	200 Westboro Road
Sample_contact_city	North Grafton
Sample_contact_state	MA
Sample_contact_zip/postal_code	01536
Sample_contact_country	USA
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM400nnn/GSM400507/suppl/GSM400507.CEL.gz
Sample_series_id	GSE15999
Sample_data_row_count	45101

GSM400508

GPL1261

Pooled lung tissue (n=8) 7 days after pneumonectomy

Lung tissue, 7 days after left unilateral pneumonectomy

strain: C57BL/6 gender: Female age: 10-12 weeks treatment: left unilateral pneumonectomy time: 7 days

Gene expression data from 8 pooled lung tissue samples 7 days after left unilateral pneumonectomy

Sample_geo_accession	GSM400508
Sample_status	Public on May 08 2009
Sample_submission_date	May 06 2009
Sample_last_update_date	May 07 2009
Sample_type	RNA
Sample_channel_count	1
Sample_organism_ch1	Mus musculus
Sample_taxid_ch1	10090
Sample_treatment_protocol_ch1	Mice were anesthetized by intraperitoneal injection of ketamine (50-75 mg/kg) and xylazine (5 mg/kg), and then received 2ml of warmed normal saline and 100mg/kg sodium ampicillin subcutaneously. Orotracheal intubation was performed under direct visualization using a 20-gauge catheter (BD Insyte catheter; Becton, Dickinson and Co, Franklin Lakes NJ) over a flexible stylet. Mice were secured in supine position, and mechanically ventilated (AUT6110, Buxco Electronics, Wilmington, NC) at 200 tidal breaths of 0.3 ml of room air per minute, at positive end-expiratory pressure of 3 cm H2O during surgery and recovery. After achieving adequate anesthetic depth (absence of response to toe-pinch) the left thoracic wall was clipped and disinfected. The skin, chest wall and pleura were incised at the 5th intercostal space, and the left lung was gently lifted through a ~5-7mm incision and ligated at the hilum with 4-0 silk (Sofsilk, Synture Norwalk Ct). The lungs were then inflated to 30 cmH20 airway pressure, and the chest wall closed during this inflation with a single interrupted suture. The skin was closed with 5-0 PDS in a simple interrupted pattern. Mice were extubated at the onset of vigorous spontaneous breathing. The mice recovered from surgery in a warmed cage, and post-operative pain was managed with buprenorphine subcutaneously (0.05mg/kg) as soon as mice showed conscious motor control, and every 12 hours thereafter as needed (<3 days). Chow, nutrient gel (on the cage floor), and water were provided ad libitum. Sham pneumonectomy animals underwent an identical procedure, except that after the thoracotomy, the chest was left open for 5 minutes to simulate the conditions of the pneumonectomy group without removal of the left lung, then closed as described.
Sample_growth_protocol_ch1	Mice used in this study were adult (10-12 week) female C57BL/6 (20-25g) obtained from Jackson Laboratories.
Sample_molecule_ch1	total RNA
Sample_extract_protocol_ch1	The mice were anesthetized as above at 6 hours, 1 day, 3 days and 7 days after surgery (PNY or SHAM) then euthanized by cervical dislocation. The pulmonary vasculature was perfused with ice cold Hanks balanced salt solution, the trachea cannulated, and the lungs removed en bloc. RNA preservation was achieved by flooding the lung intratracheally with RNAlater solution (Qiagen #76104), followed by storage of lung tissue samples in RNAlater at -80ºC. Equal amounts of lung tissue were pooled from eight animals in each group (PNY or SHAM at each of 4 time points) to minimize biological variability. Total RNA was prepared from the dissected lung tissue using the Qiagen RNAeasy mini kit (Qiagen #74104) according to the manufacturer’s directions. The total RNA samples from the primary purification were purified a second time on Qiagen RNAeasy columns according to the manufacturer’s instructions. Total RNA concentrations, A260/A280 and A260/A230 ratios was determined using a NanoDrop ND1000 spectrophotometer.
Sample_label_ch1	Biotin
Sample_label_protocol_ch1	Labeling was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_hyb_protocol	Chip hybridization was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_scan_protocol	GeneChips were scanned using the Affymetrix 7G scanner.
Sample_data_processing = The initial goal in the analysis was to identify genes regulated in the comparison between pneumonectomized and sham-operated animals. By treating all time points as replicates within their respective groups two datasets were created (PNY and SHAM), each with an n	4. The corresponding Affymetrix CEL files were background corrected, summarized and quantile-normalized using the RMA library in BioConductor (http:www.bioconductor.org), yielding one expression value per probe set for each of the 8 arrays. Based on the ‘Rank Products’ algorithm proposed by Brietling, et. al, the RankProd library was employed to find differentially expressed genes. This algorithm works by performing comprehensive pair-wise comparisons to calculate a rank statistic RPg, the probability of seeing the observed, pair-wise expression patterns for any given gene g. As a vehicle for measuring statistical significance a non-parametric P-value is also calculated, using 1000 permutations to determine how often the calculated RPg statistic would occur by chance alone. Finally, the RankProd library compares average expression between the two groups to derive a fold-change value. Genes with a reported P-value < 0.001 and a fold-change > 1.5 or < -1.5 were selected for further investigation. In a second analysis, the focus was shifted to temporal changes in gene expression during lung regeneration as opposed to overall transcriptomic patterns. With only one array per experimental condition at each time point, derivation of statistical measures and the subsequent search for truly differentially expressed genes can be challenging. However, the S-Score algorithm described by Zhang et al. (2002) and Kerns et al. (2003) provides a method for determining statistical significance when biological replicates are not available by applying pairwise comparisons to probe-level data. On average, the Affymetrix 3’ IVT platform contains 22 probes for every transcript represented on the array. Using this information directly, the S-Score algorithm has shown an increased sensitivity when compared to many other existing analysis methods without sacrificing specificity, and can produce accurate results when no biological replicates are present (This is particularly applicable and appropriate to our individual time point datasets in which we have only one paired array set for each time point. Using the S-Score algorithm, the relative change in probe pair intensity is calculated to convert the probe pair signal differences into multiple measurements with equalized errors. The relative changes for each probe pair are then summed to form the S-score, which represents a single measure of the significance of change for the gene in question. By definition, S-score is related to P-value by an exponential relation, and a value of 3 corresponds to a P-value of 0.003 . Therefore, genes with an S-score > 3.0 or < - 3.0 (P < 0.003) were selected for further analysis.
Sample_platform_id	GPL1261
Sample_contact_name	Julia,A,Paxson
Sample_contact_email	julia.paxson@tufts.edu
Sample_contact_laboratory	Hoffman Lab
Sample_contact_department	Clinical Sciences
Sample_contact_institute	Tufts Cummings School of Veterinary Medicine
Sample_contact_address	200 Westboro Road
Sample_contact_city	North Grafton
Sample_contact_state	MA
Sample_contact_zip/postal_code	01536
Sample_contact_country	USA
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM400nnn/GSM400508/suppl/GSM400508.CEL.gz
Sample_series_id	GSE15999
Sample_data_row_count	45101

GSM400509

GPL1261

Pooled lung tissue (n=8) 7 days after sham thoracotomy

Lung tissue, 7 days after sham thoracotomy

strain: C57BL/6 gender: Female age: 10-12 weeks treatment: sham thoracotomy time: 7 days

Gene expression data from 8 pooled lung tissue samples 7 days after sham thoracotomy

Sample_geo_accession	GSM400509
Sample_status	Public on May 08 2009
Sample_submission_date	May 06 2009
Sample_last_update_date	May 07 2009
Sample_type	RNA
Sample_channel_count	1
Sample_organism_ch1	Mus musculus
Sample_taxid_ch1	10090
Sample_treatment_protocol_ch1	Mice were anesthetized by intraperitoneal injection of ketamine (50-75 mg/kg) and xylazine (5 mg/kg), and then received 2ml of warmed normal saline and 100mg/kg sodium ampicillin subcutaneously. Orotracheal intubation was performed under direct visualization using a 20-gauge catheter (BD Insyte catheter; Becton, Dickinson and Co, Franklin Lakes NJ) over a flexible stylet. Mice were secured in supine position, and mechanically ventilated (AUT6110, Buxco Electronics, Wilmington, NC) at 200 tidal breaths of 0.3 ml of room air per minute, at positive end-expiratory pressure of 3 cm H2O during surgery and recovery. After achieving adequate anesthetic depth (absence of response to toe-pinch) the left thoracic wall was clipped and disinfected. The skin, chest wall and pleura were incised at the 5th intercostal space, and the left lung was gently lifted through a ~5-7mm incision and ligated at the hilum with 4-0 silk (Sofsilk, Synture Norwalk Ct). The lungs were then inflated to 30 cmH20 airway pressure, and the chest wall closed during this inflation with a single interrupted suture. The skin was closed with 5-0 PDS in a simple interrupted pattern. Mice were extubated at the onset of vigorous spontaneous breathing. The mice recovered from surgery in a warmed cage, and post-operative pain was managed with buprenorphine subcutaneously (0.05mg/kg) as soon as mice showed conscious motor control, and every 12 hours thereafter as needed (<3 days). Chow, nutrient gel (on the cage floor), and water were provided ad libitum. Sham pneumonectomy animals underwent an identical procedure, except that after the thoracotomy, the chest was left open for 5 minutes to simulate the conditions of the pneumonectomy group without removal of the left lung, then closed as described.
Sample_growth_protocol_ch1	Mice used in this study were adult (10-12 week) female C57BL/6 (20-25g) obtained from Jackson Laboratories.
Sample_molecule_ch1	total RNA
Sample_extract_protocol_ch1	The mice were anesthetized as above at 6 hours, 1 day, 3 days and 7 days after surgery (PNY or SHAM) then euthanized by cervical dislocation. The pulmonary vasculature was perfused with ice cold Hanks balanced salt solution, the trachea cannulated, and the lungs removed en bloc. RNA preservation was achieved by flooding the lung intratracheally with RNAlater solution (Qiagen #76104), followed by storage of lung tissue samples in RNAlater at -80ºC. Equal amounts of lung tissue were pooled from eight animals in each group (PNY or SHAM at each of 4 time points) to minimize biological variability. Total RNA was prepared from the dissected lung tissue using the Qiagen RNAeasy mini kit (Qiagen #74104) according to the manufacturer’s directions. The total RNA samples from the primary purification were purified a second time on Qiagen RNAeasy columns according to the manufacturer’s instructions. Total RNA concentrations, A260/A280 and A260/A230 ratios was determined using a NanoDrop ND1000 spectrophotometer.
Sample_label_ch1	Biotin
Sample_label_protocol_ch1	Labeling was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_hyb_protocol	Chip hybridization was performed as described in the Affymetrix GeneChip Expression Analysis Technical Manual using Affymetrix Mouse Genome 430 2.0 GeneChips and the One-Cycle cDNA Synthesis and HT IVT Labeling kits (Affymetrix Inc.).
Sample_scan_protocol	GeneChips were scanned using the Affymetrix 7G scanner.
Sample_data_processing = The initial goal in the analysis was to identify genes regulated in the comparison between pneumonectomized and sham-operated animals. By treating all time points as replicates within their respective groups two datasets were created (PNY and SHAM), each with an n	4. The corresponding Affymetrix CEL files were background corrected, summarized and quantile-normalized using the RMA library in BioConductor (http:www.bioconductor.org), yielding one expression value per probe set for each of the 8 arrays. Based on the ‘Rank Products’ algorithm proposed by Brietling, et. al, the RankProd library was employed to find differentially expressed genes. This algorithm works by performing comprehensive pair-wise comparisons to calculate a rank statistic RPg, the probability of seeing the observed, pair-wise expression patterns for any given gene g. As a vehicle for measuring statistical significance a non-parametric P-value is also calculated, using 1000 permutations to determine how often the calculated RPg statistic would occur by chance alone. Finally, the RankProd library compares average expression between the two groups to derive a fold-change value. Genes with a reported P-value < 0.001 and a fold-change > 1.5 or < -1.5 were selected for further investigation. In a second analysis, the focus was shifted to temporal changes in gene expression during lung regeneration as opposed to overall transcriptomic patterns. With only one array per experimental condition at each time point, derivation of statistical measures and the subsequent search for truly differentially expressed genes can be challenging. However, the S-Score algorithm described by Zhang et al. (2002) and Kerns et al. (2003) provides a method for determining statistical significance when biological replicates are not available by applying pairwise comparisons to probe-level data. On average, the Affymetrix 3’ IVT platform contains 22 probes for every transcript represented on the array. Using this information directly, the S-Score algorithm has shown an increased sensitivity when compared to many other existing analysis methods without sacrificing specificity, and can produce accurate results when no biological replicates are present (This is particularly applicable and appropriate to our individual time point datasets in which we have only one paired array set for each time point. Using the S-Score algorithm, the relative change in probe pair intensity is calculated to convert the probe pair signal differences into multiple measurements with equalized errors. The relative changes for each probe pair are then summed to form the S-score, which represents a single measure of the significance of change for the gene in question. By definition, S-score is related to P-value by an exponential relation, and a value of 3 corresponds to a P-value of 0.003 . Therefore, genes with an S-score > 3.0 or < - 3.0 (P < 0.003) were selected for further analysis.
Sample_platform_id	GPL1261
Sample_contact_name	Julia,A,Paxson
Sample_contact_email	julia.paxson@tufts.edu
Sample_contact_laboratory	Hoffman Lab
Sample_contact_department	Clinical Sciences
Sample_contact_institute	Tufts Cummings School of Veterinary Medicine
Sample_contact_address	200 Westboro Road
Sample_contact_city	North Grafton
Sample_contact_state	MA
Sample_contact_zip/postal_code	01536
Sample_contact_country	USA
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM400nnn/GSM400509/suppl/GSM400509.CEL.gz
Sample_series_id	GSE15999
Sample_data_row_count	45101

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