Journal: The American journal of pathology

Article Title: Pseudogene-Derived Long Noncoding RNAs GSTM3P1/Gstm2-ps1 Exacerbate Sepsis-Associated Acute Kidney Injury by Suppressing Their Parent Gene Translation

doi: 10.1016/j.ajpath.2026.01.004

Figure Lengend Snippet: GSTM3P1/Gstm2-ps1 are transiently induced in kidney proximal tubular cells in sepsis-associated acute kidney injury models. BUMPT and HK-2 cells were treated with 100 μg/mL lipopolysaccharide (LPS) for 3, 6, and 12 hours. Male C57BL/6J mice were subjected to an i.p. injection of 10 mg/kg LPS or cecal ligation and puncture (CLP) surgery, and kidney samples were collected at 3, 6, and 9 hours after injury. A—D: Quantitative RT-PCR analysis of Gstm2-ps1 or GSTM3P1 expression. A: Gstm2-ps1 in BUMPT. B: GSTM3P1 in HK2. C: Gstm2-ps1 in LPS-treated mouse kidney. D: Gstm2-ps1 in mouse kidney with CLP. A—D: The statistical difference was determined using one-way analysis of variance with Tukey multiple comparison. E and F: Representative images of Gstm2-ps1 in kidney tissue assessed by in situ hybridization and costained with fluorescein isothiocyanate—labeled Lotus tetragonolobus lectin (LTL). E: Mouse kidneys with or without LPS treatment for 3 hours. F: Mouse kidneys with or without CLP injury for 3 hours. n 5 ( A—D ); n 6 ( E and F ). ** P < 0.01, *** P < 0.001. Scale bar 100 μm ( E and F ). NS, nonsignificant.

Article Snippet: Human GSTM3 or murine Gstm2 open reading frames were subcloned from commercially available plasmids (RC201013 or MR202469; Origene, Rockville, MD), respectively.

Techniques: Injection, Ligation, Quantitative RT-PCR, Expressing, Comparison, In Situ Hybridization, Labeling

Journal: The American journal of pathology

Article Title: Pseudogene-Derived Long Noncoding RNAs GSTM3P1/Gstm2-ps1 Exacerbate Sepsis-Associated Acute Kidney Injury by Suppressing Their Parent Gene Translation

doi: 10.1016/j.ajpath.2026.01.004

Figure Lengend Snippet: GSTM3P1/Gstm2-ps1 exacerbate lipopolysaccharide (LPS)—induced apoptosis in cultured proximal tubular cells. RPTC cells with or without GSTM3P1 stable overexpression, BUMPT cells with or without Gstm2-ps1 stable overexpression, and wild-type (WT) or Gstm2-ps1 knockout (KO) primary proximal tubular cells were treated with 100 μg/mL LPS for 19 hours. A—C: Quantitative RT-PCR analysis confirming the expression of GSTM3P1 or Gstm2-ps1. A: RPTC. B: BUMPT. C: Primary proximal tubular cells. D—F: Cell viability assessed by MTT assay following 19 hours of LPS treatment. D: RPTC. E: BUMPT. F: Primary proximal tubular cells. A—F: The statistical difference was determined by unpaired t -test. G—I: Representative immunoblots of full-length caspase 3 and cleaved caspase 3 (C-caspase-3) with β-actin as the loading control (Ctrl; top panels ), and the densitometry analysis of C-caspase-3 normalized to β-actin ( bottom panels ). G: RPTC. H: BUMPT. I: Primary proximal tubular cells. G—I: The statistical difference was determined by two-way analysis of variance with Tukey multiple comparison. n =3 ( A , G , and H ); n = 4 ( B , C , and I ); n = 6 ( D—F ). * P < 0.05, ** P < 0.01, and *** P < 0.001.

Article Snippet: Human GSTM3 or murine Gstm2 open reading frames were subcloned from commercially available plasmids (RC201013 or MR202469; Origene, Rockville, MD), respectively.

Techniques: Cell Culture, Over Expression, Knock-Out, Quantitative RT-PCR, Expressing, MTT Assay, Western Blot, Control, Comparison

Journal: The American journal of pathology

Article Title: Pseudogene-Derived Long Noncoding RNAs GSTM3P1/Gstm2-ps1 Exacerbate Sepsis-Associated Acute Kidney Injury by Suppressing Their Parent Gene Translation

doi: 10.1016/j.ajpath.2026.01.004

Figure Lengend Snippet: GSTM3P1/Gstm2-ps1 overexpression induces oxidative stress in cultured proximal tubular cells. RPTC cells with or without GSTM3P1 stable overexpression and BUMPT cells with or without Gstm2-ps1 stable overexpression were treated with 100 μg/mL lipopolysaccharide (LPS) for 19 hours. A and B: Oxidative stress levels were assessed by measuring the reduced glutathione (GSH)/oxidized glutathione (GSSG) ratio. A: RPTC. B: BUMPT. The statistical difference was determined by two-way analysis of variance with Tukey multiple comparison. C: Representative images of 2′,7′-dichlorodihydrofluorescein diacetate (DCFH-DA)—stained RPTC and phase-contrast images. n = 6 ( A—C ). * P < 0.05, ** P < 0.01, and *** P < 0.001. Scale bars = 200 μm ( C ).

Article Snippet: Human GSTM3 or murine Gstm2 open reading frames were subcloned from commercially available plasmids (RC201013 or MR202469; Origene, Rockville, MD), respectively.

Techniques: Over Expression, Cell Culture, Comparison, Staining

Journal: The American journal of pathology

Article Title: Pseudogene-Derived Long Noncoding RNAs GSTM3P1/Gstm2-ps1 Exacerbate Sepsis-Associated Acute Kidney Injury by Suppressing Their Parent Gene Translation

doi: 10.1016/j.ajpath.2026.01.004

Figure Lengend Snippet: Proximal tubular—specific Gstm2-ps1 knockout protects mice from lipopolysaccharide (LPS)—induced sepsis acute kidney injury. Proximal tubular—specific Gstm2-ps1 knockout (KO) mice and their wild-type (WT) littermates were treated with one i.p. injection of 10 mg/kg LPS and sacrificed after 19 hours. The control (Ctrl) mice were given an equal volume of phosphate-buffered saline. A: Blood urea nitrogen (BUN) levels. B: Serum creatinine levels. C: Quantification of apoptotic cells in kidneys by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining. A—C: The statistical difference was determined by two-way analysis of variance with Tukey multiple comparison. D: Representative images of TUNEL staining. E: Representative images of neutrophil gelatinase-associated lipocalin (NGAL) immunohistochemical staining. F: Representative images of reactive oxygen species (ROS) levels detected by 2′,7′-dichlorodihydrofluorescein diacetate staining. G: Representative immunoblots of kidney injury molecule-1 (KIM-1) and the densitometry analysis of KIM-1 normalized to the internal loading control glyceraldehyde-3-phosphate dehydrogenase. Data are presented as means ± SD ( A—C and G ). n = 6 per group ( A , B , E , and F ); n = 5 per group ( C ); n = 4 per group ( G ). ** P < 0.01, *** P < 0.001, and **** P < 0.0001. Scale bars: 50 μm ( D and E ); 100 μm ( F ).

Article Snippet: Human GSTM3 or murine Gstm2 open reading frames were subcloned from commercially available plasmids (RC201013 or MR202469; Origene, Rockville, MD), respectively.

Techniques: Knock-Out, Injection, Control, Saline, End Labeling, TUNEL Assay, Staining, Comparison, Immunohistochemical staining, Western Blot

Journal: The American journal of pathology

Article Title: Pseudogene-Derived Long Noncoding RNAs GSTM3P1/Gstm2-ps1 Exacerbate Sepsis-Associated Acute Kidney Injury by Suppressing Their Parent Gene Translation

doi: 10.1016/j.ajpath.2026.01.004

Figure Lengend Snippet: GSTM3P1/Gstm2-ps1 down-regulate the expression of their parent gene GSTM3/GSTM2. A—F: RPTC cells ( A and D ) or BUMPT cells ( B and E ) were overexpressed with or without GSTM3P1 or Gstm2-ps1, and C57BL/6J mouse kidneys ( C and F ) were treated with 10 mg/kg lipopolysaccharide (LPS) for 3, 6, and 9 hours. A—C: Representative immunoblots of GSTM3 and GSTM2, with β-actin or cyclophilin B as the internal loading marker. D—F: The densitometric analysis of GSTM3 and GSTM2 protein normalized to the internal loading control. The statistical difference was determined by unpaired t -test ( D and E ) or one-way analysis of variance with Tukey multiple comparison ( F ). G: Representative immunofluorescence images of GSTM2 with fluorescein isothiocyanate— Lotus tetragonolobus lectin (LTL) costaining. Gstm2-ps1 knockout (KO) and wild-type (WT) mice were treated with/without 10 mg/kg LPS for 3 hours. n = 6 for cell samples ( D—F ); n = 4 for kidney samples ( D—F ); n = 6 ( G ). ** P < 0.01, *** P < 0.001, and **** P < 0.0001. Scale bar = 100 μm ( G ). NS, nonsignificant.

Article Snippet: Human GSTM3 or murine Gstm2 open reading frames were subcloned from commercially available plasmids (RC201013 or MR202469; Origene, Rockville, MD), respectively.

Techniques: Expressing, Western Blot, Marker, Control, Comparison, Immunofluorescence, Knock-Out

Journal: The American journal of pathology

Article Title: Pseudogene-Derived Long Noncoding RNAs GSTM3P1/Gstm2-ps1 Exacerbate Sepsis-Associated Acute Kidney Injury by Suppressing Their Parent Gene Translation

doi: 10.1016/j.ajpath.2026.01.004

Figure Lengend Snippet: GSTM2/GSTM3 protect cultured renal proximal tubular cell from lipopolysaccharide (LPS)—induced apoptosis. RPTC cells with or without GSTM3 overexpression, and BUMPT cells with or without GSTM2 overexpression, were treated with 100 μg/mL LPS. A and B: Immunoblot analysis confirming the overexpression efficiency of GSTM3 and GSTM2. C and D: Cell viability assessed using the MTT assay following 17 hours of LPS treatment. A—D: The statistical difference was determined by unpaired t -test. E and F: Representative immunoblots ( top panels ) and densitometric analysis ( bottom panels ) of cleaved caspase-3 (C-caspase-3) protein levels. E: RPTC. F: BUMPT. E and F: The statistical difference was determined by one-way analysis of variance with Tukey multiple comparison. n = 3 ( A and B ); n = 6 ( C and D ); n = 4 ( E and F ). ** P < 0.01, *** P < 0.001, and **** P < 0.0001. Ctrl, control.

Article Snippet: Human GSTM3 or murine Gstm2 open reading frames were subcloned from commercially available plasmids (RC201013 or MR202469; Origene, Rockville, MD), respectively.

Techniques: Cell Culture, Over Expression, Western Blot, MTT Assay, Comparison, Control

Journal: The American journal of pathology

Article Title: Pseudogene-Derived Long Noncoding RNAs GSTM3P1/Gstm2-ps1 Exacerbate Sepsis-Associated Acute Kidney Injury by Suppressing Their Parent Gene Translation

doi: 10.1016/j.ajpath.2026.01.004

Figure Lengend Snippet: GSTM3P1/Gstm2-ps1 may down-regulate GSTM3/GSTM2 protein level through RNA translation inhibition. A: Quantitative RT-PCR (RT-qPCR) of GSTM3 in RPTC cells with or without GSTM3P1 overexpression. The unpaired t -test was used. B: RT-qPCR of GSTM2 in BUMPT cells with or without Gstm2-ps1 overexpression. The unpaired t -test was used. C: RT-qPCR of GSTM2 in mouse kidneys with or without 10 mg/kg lipopolysaccharide (LPS) treatment . The one-way analysis of variance (ANOVA) with Tukey multiple comparison was used. D and E: BUMPT cell lysates with Gstm2-ps1 overexpression or empty vector transfection (pcDNA) were subjected to RNA pull-down assay with Gstm2-ps1 probes. The input lysates (Input) and pull-down products (pull-down) were examined by immunoblotting of human antigen R (HuR), with cyclophilin B as the internal loading control of input lysates. D: Representative immunoblots of HuR and cyclophilin B. E: The densitometry analysis of HuR immunoblotting in pull-down products normalized by its level in the input. The unpaired t -test was used. F—H: BUMPT cells were overexpressed with HuR or Gstm2-ps1. F: Representative immunoblots to confirm the overexpression of HuR with cyclophilin B as the internal loading control. The upper band is the overexpressed HuR. The lower band is the endogenous HuR. G: Representative immunoblots and densitometry analysis of GSTM2 in cells with or without Gstm2-ps1 and HuR overexpression. β-Actin was used as the internal loading control. The one-way ANOVA with Tukey multiple comparison was used. H: The cell viability assessed by MTT assay. The one-way ANOVA with Tukey multiple comparison was used. n = 5 ( A and B ); n = 4 ( C and E—G ); n = 6 ( H ). * P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001. NS, nonsignificant.

Article Snippet: Human GSTM3 or murine Gstm2 open reading frames were subcloned from commercially available plasmids (RC201013 or MR202469; Origene, Rockville, MD), respectively.

Techniques: Inhibition, Quantitative RT-PCR, Over Expression, Comparison, Plasmid Preparation, Transfection, Pull Down Assay, Western Blot, Control, MTT Assay

Journal: Cancers

Article Title: An Inducible BRCA1 Expression System with In Vivo Applicability Uncovers Activity of the Combination of ATR and PARP Inhibitors to Overcome Therapy Resistance

doi: 10.3390/cancers18020309

Figure Lengend Snippet: ( A ) ( Left panel ): dose–response curves of olaparib in colony formation assays in tetracycline-repressor (TR) expressing MDAMB436 cells with or without RAD51B complementation (MDAMB436-B-TR and MDAMB436-TR, respectively) and with (+BRCA1) or without (+LacZ) BRCA1 complementation. ( Right panel ): Logarithmic half-maximal inhibitory concentration (LogIC50) of olaparib for each cell line. ( B ) ( Left panel ) : dose–response curves of olaparib in colony formation assays in MDAMB436-B-TR cells expressing full-length BRCA1 (+FL), no BRCA1 (+LacZ), or different BRCA1 hypomorphs (+RING-less, +∆exon11, +BRCT-less). ( Right panel ): Logarithmic half-maximal inhibitory concentration (LogIC50) of olaparib for each cell line. ( C ) ( Left panel ): dose–response curves of olaparib in survival assays in MDAMB436-B-TR cells expressing full-length BRCA1 (+FL), no BRCA1 (+LacZ), or different BRCA1 hypomorphs (+C64R, +L1407P, +R1699Q). ( Right panel ): Logarithmic half-maximal inhibitory concentration (LogIC50) of olaparib for each cell line. Data for BRCA1 hypomorphs with deletions in the RING (RING-less, M48 START) or BRCT (BRCT-less) domains are included for comparative purposes. ( D ) ( Left panel ): dose–response curves of carboplatin in survival assays in MDAMB436-B-TR cells expressing full-length BRCA1 (+FL), no BRCA1 (+LacZ) or different BRCA1 hypomorphs (+RING-less, +∆exon11, +BRCT-less, +CC-mutant). ( Right panel ): Logarithmic half-maximal inhibitory concentration (LogIC50) of carboplatin for each cell line. ( E ) Western blot of immunoprecipitation experiments in MDAMB436-B-TR cells expressing full-length (+FL) or ∆exon11 BRCA1 and exposed to different doxycycline doses. GAPDH was used as a loading control. I = input; F = FLAG immunoprecipitation. ( F ) Logarithmic half-maximal inhibitory concentration (LogIC50) of olaparib at the different doses of doxycycline used. All data are from at least 3 biological replicates. Statistical analysis performed using One-Way ANOVA with Holm–Sidak multiple comparisons, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. CFU = colony-forming units. Dotted lines in the olaparib IC50 graphs represent the minimal free concentration of olaparib (approximately 300 nM) in the plasma of patients on the established monotherapy dose of 300 mg twice daily. The original western blot figures can be found in .

Article Snippet: Plasmids containing the human BRCA1 (NM_007294) and RAD51B (NM_133509) open reading frames (ORFs) were obtained from Genscript (Ohu18572D) and OriGene (RC206457L3), respectively.

Techniques: Expressing, Concentration Assay, Mutagenesis, Western Blot, Immunoprecipitation, Control, Clinical Proteomics