TRPC6 Knockout Alleviates Renal Fibrosis through PI3K/AKT/GSK3B Pathway.

OBJECTIVE: Renal fibrosis is the ultimate pathway of various forms of acute and chronic kidney damage. Notably, the knockout of transient receptor potential channel 6 (TRPC6) has shown promise in alleviating renal fibrosis. However, the regulatory impact of TRPC6 on renal fibrosis remains unclear. METHODS: In vivo, TRPC6 knockout (TRPC6-/-) mice and age-matched 129 SvEv (WT) mice underwent unilateral renal ischemia-reperfusion (uIR) injury surgery on the left renal pedicle or sham operation. Kidneys and serum were collected on days 7, 14, 21, and 28 after euthanasia. In vitro, primary tubular epithelial cells (PTECs) were isolated from TRPC6-/- and WT mice, followed by treatment with transforming growth factor ß1 (TGFß1) for 72 h. The anti-fibrotic effect of TRPC6-/- and the underlying mechanisms were assessed through hematoxylin-eosin staining, Masson staining, immunostaining, qRT-PCR, and Western blotting. RESULTS: Increased TRPC6 expression was observed in uIR mice and PTECs treated with TGFß1. TRPC6-/- alleviated renal fibrosis by reducing the expression of fibrotic markers (Col-1, α-SMA, and vimentin), as well as decreasing the apoptosis and inflammation of PTECs during fibrotic progression both in vivo and in vitro. Additionally, we found that the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT)/glycogen synthase kinase 3 beta (GSK3ß) signaling pathway, a pivotal player in renal fibrosis, was down-regulated following TRPC6 deletion. CONCLUSION: These results suggest that the ablation of TRPC6 may mitigate renal fibrosis by inhibiting the apoptosis and inflammation of PTECs through down-regulation of the PI3K/AKT/GSK3ß pathway. Targeting TRPC6 could be a novel therapeutic strategy for preventing chronic kidney disease.

Apigenin suppresses epithelial-mesenchymal transition in high glucose-induced retinal pigment epithelial cell by inhibiting CBP/p300-mediated histone acetylation.

Epithelial mesenchymal transition (EMT) is a critical process implicated in the pathogenesis of retinal fibrosis and the exacerbation of diabetic retinopathy (DR) within retinal pigment epithelium (RPE) cells. Apigenin (AP), a potential dietary supplement for managing diabetes and its associated complications, has demonstrated inhibitory effects on EMT in various diseases. However, the specific impact and underlying mechanisms of AP on EMT in RPE cells remain poorly understood. In this study, we have successfully validated the inhibitory effects of AP on high glucose-induced EMT in ARPE-19 cells and diabetic db/db mice. Notably, our findings have identified CBP/p300 as a potential therapeutic target for EMT in RPE cells and have further substantiated that AP effectively downregulates the expression of EMT-related genes by attenuating the activity of CBP/p300, consequently reducing histone acetylation alterations within the promoter region of these genes. Taken together, our results provide novel evidence supporting the inhibitory effect of AP on EMT in RPE cells, and highlight the potential of specifically targeting CBP/p300 as a strategy for inhibiting retinal fibrosis in the context of DR.

Identification of ALDOB as a novel prognostic biomarker in kidney clear cell renal cell carcinoma.

Kidney clear cell renal cell carcinoma (KIRC) is also the most lethal subtype among all kidney cancer subtypes, posing a severe threat to public health. Therefore, it is crucial to identify new, reliable biomarkers in KIRC. Therefore, it is crucial to identify novel, reliable biomarkers associated with KIRC. We analyzed RNA sequence results from TCGA and several GEO datasets. The commonly deregulated gene, ALDOB, was found in multiple data and confirmed its important prognostic value. Subsequently, we explored the specific mechanism by which ALDOB regulates anti-tumor immunity through in vivo and in vitro experiments. We found that ALDOB may play a role in regulating tumor growth by regulating CD8+ T cell infiltration. This is consistent with the results of our immune infiltration-related analysis. In addition, we have also discovered the effect of ALDOB in previous studies on other cancer types. Finally, we concluded that ALDOB may have potential reference value for immunotherapy and can also be used as an independent predictor of prognosis in KIRC.

Loureirin B ameliorates cholestatic liver fibrosis via AKT/mTOR/ATG7-mediated autophagy of hepatic stellate cells.

AIM OF THE STUDY: Chronic cholestasis leads to liver fibrosis, which lacks effective treatment. In this study, we investigated the role and mechanisms of action of loureirin B (LB) in cholestatic liver fibrosis. MATERIALS AND METHODS: Bile duct ligation (BDL)-induced hepatic fibrosis mice were used as in vivo models. Transforming growth factor-ß1 (TGF-ß1)-pretreated HSC-T6 cells were used to explore the mechanism by which LB attenuates liver fibrosis in vitro. RNA sequencing, quantitative PCR (qPCR), western blotting, immunohistochemistry and immunofluorescence were performed to detect the fibrosis markers and measure autophagy levels. Flow cytometry, cell counting kit-8 (CCK-8) assay, and 5'-ethynyl-2'-deoxyuridine (EdU) assay were conducted to detect cell proliferation and viability. GFP-RFP-LC3 adenovirus, autophagy-related protein 7 (ATG7) siRNA, and bafilomycin A1 (BafA1) were used to verify autophagic flux. RESULTS: Our results showed that LB ameliorates liver injury, inhibits collagen deposition, and decreases the expressions of fibrosis-related markers in BDL-induced mouse livers. In vitro, we found that LB inhibited proliferation and migration, promoted apoptosis, and inhibited the activation of HSC-T6 cells pretreated with TGF-ß1. RNA sequencing analysis of HSC-T6 cells showed that LB treatment predominantly targeted autophagy-related pathways. Further protein analysis indicated that LB downregulated the expression of phosphorylated AKT (p-AKT) and phosphorylated mTOR (p-mTOR), and upregulated LC3-II, p62, and ATG7 both in vivo and in vitro. Intriguingly, ATG7 inactivation reversed the antifibrotic effects of LB on HSC-T6 cells. CONCLUSIONS: LB can improve BDL-induced liver fibrosis by inhibiting the activation and proliferation of HSCs and is expected to be a promising antifibrotic drug.

Paeoniflorin promotes PPARγ expression to suppress HSCs activation by inhibiting EZH2-mediated histone H3K27 trimethylation.

BACKGROUND: The alleviating effect of paeoniflorin (Pae) on liver fibrosis has been established; however, the molecular mechanism and specific target(s) underlying this effect remain elusive. PURPOSE: This study was to investigate the molecular mechanism underlying the regulatory effect of Pae on hepatic stellate cells (HSCs) activation in liver fibrosis, with a specific focus on the role of Pae in modulating histone methylation modifications. METHODS: The therapeutic effect of Pae was evaluated by establishing in vivo and in vitro models of carbon tetrachloride (CCl4)-induced mice and transforming growth factor ß1 (TGF-ß1)-induced LX-2 cells, respectively. Molecular docking, surface plasmon resonance (SPR), chromatin immunoprecipitation-quantitative real time PCR (ChIP-qPCR) and other molecular biological methods were used to clarify the molecular mechanism of Pae regulating HSCs activation. RESULTS: Our study found that Pae inhibited HSCs activation and histone trimethylation modification in liver of CCl4-induced mice and LX-2 cells. We demonstrated that the inhibitory effect of Pae on the activation of HSCs was dependent on peroxisome proliferator-activated receptor γ (PPARγ) expression and enhancer of zeste homolog 2 (EZH2). Mechanistically, Pae directly binded to EZH2 to effectively suppress its enzymatic activity. This attenuation leaded to the suppression of histone H3K27 trimethylation in the PPARγ promoter region, which induced upregulation of PPARγ expression. CONCLUSION: This investigative not only sheds new light on the precise targets that underlie the remission of hepatic fibrogenesis induced by Pae but also emphasizes the critical significance of EZH2-mediated H3K27 trimethylation in driving the pathogenesis of liver fibrosis.

Si-Ni-San inhibits hepatic Fasn expression and lipid accumulation in MAFLD mice through AMPK/p300/SREBP-1c axis.

BACKGROUND: Soothing the liver and regulating qi is one of the core ideas of traditional Chinese medicine (TCM) in the treatment of fatty liver. Si-Ni-San (SNS) is a well-known herbal formula in TCM for liver soothing and qi regulation in fatty liver treatment. However, its efficacy lacks modern scientific evidence. PURPOSE: This study was aimed to investigate the impact of SNS on metabolic associated fatty liver disease (MAFLD) in mice and explore the underlying molecular mechanisms, particularly its effects on lipid metabolism in hepatocytes. METHODS: The therapeutic effect of SNS was evaluated using in vivo and in vitro models of high-fat/high-cholesterol (HFHC) diet-induced mice and palmitic acid (PA)-induced hepatocytes, respectively. Molecular biological techniques such as RNA-sequencing (RNA-seq), co-immunoprecipitation (co-IP), and western blotting were employed to elucidate the molecular mechanism of SNS in regulating lipid metabolism in hepatocytes. RESULTS: Our findings revealed that SNS effectively reduced lipid accumulation in the livers of HFHC diet-induced mice and PA-induced hepatocytes. RNA-seq analysis demonstrated that SNS significantly down-regulated the expression of fatty acid synthase (Fasn) in the livers of HFHC-fed mice. Mechanistically, SNS inhibited Fasn expression and lipid accumulation by activating adenosine monophosphate (AMP)-activated protein kinase (AMPK). Activation of AMPK suppressed the activity of the transcriptional coactivator p300 and modulated the protein stability of sterol regulatory element-binding protein-1c (SREBP-1c). Importantly, p300 was required for the inhibition of Fasn expression and lipid accumulation by SNS. Furthermore, SNS activated AMPK by decreasing adenosine triphosphate (ATP) production in hepatocytes. CONCLUSION: This study provided novel evidence on the regulatory mechanisms underlying the effects of SNS on Fasn expression. Our findings demonstrate, for the first time, that SNS exerts suppressive effects on Fasn expression through modulation of the AMPK/p300/SREBP-1c axis. Consequently, this regulatory pathway mitigates excessive lipid accumulation and ameliorates MAFLD in mice.

Essential oil extracted from Quzhou Aurantii Fructus prevents acute liver failure through inhibiting lipopolysaccharide-mediated inflammatory response.

Quzhou Aurantii Fructus (QAF) has a long history as a folk medicine and food for the treatment of liver diseases. While our earlier study provided evidence of hepatoprotective properties contained within the flavonoids and limonins constituents in QAF, the potential preventative effects afforded by essential oil components present within QAF remains enigmatic. In this study, we prepared Quzhou Aurantii Fructus essential oil (QAFEO) and confirmed its anti-inflammatory effects on liver inflammation through experimentation on lipopolysaccharide and D-galactosamine (LPS/D-GalN) induced acute liver failure (ALF) mouse models. Using RNA-sequence (RNA-seq) analysis, we found that QAFEO prevented ALF by systematically blunting the pathways involved in response to LPS and toll-like receptor signaling pathways. QAFEO effectively suppressed the phosphorylation of tank-binding kinase 1 (TBK1), TGF-beta activated kinase 1 (TAK1), interferon regulatory factor 3 (IRF3), and the activation of mitogen activated kinase-like protein (MAPK) and nuclear factor-kappa B (NF-κB) pathways in vivo and in vitro. Importantly, QAFEO substantially reduced myeloid differentiation primary response gene 88 (MyD88)- toll-like receptor 4 (TLR4) interaction levels. Moreover, 8 compounds from QAFEO could directly bind to REAL, TAK1, MyD88, TBK1, and IRF3. Taken together, the results of our study support the notion that QAFEO exerts a hepatoprotective effect through inhibiting LPS-mediated inflammatory response.

Saikosaponin A triggers cell ferroptosis in hepatocellular carcinoma by inducing endoplasmic reticulum stress-stimulated ATF3 expression.

Ferroptosis is a type of nonapoptotic necrotic cell death characterized by iron-dependent lipid peroxidation. Saikosaponin A (SsA), a natural bioactive triterpenoid saponin extracted from Radix Bupleuri, has shown potent antitumor activity against various tumors. However, the underlying mechanism of the antitumor activity of SsA remains unclear. Here, we discovered that SsA induced HCC cell ferroptosis in vitro and in vivo. Using RNA-sequence analysis, we found that SsA mainly affected the glutathione metabolic pathway and inhibited the expression of cystine transporter solute carrier family 7 member 11 (SLC7A11). Indeed, SsA increased intracellular malondialdehyde (MDA) and iron accumulation, while it decreased the levels of reduced glutathione (GSH) in HCC. Deferoxamine (DFO), ferrostatin-1 (Fer-1) and GSH could rescue SsA-induced cell death, whereas Z-VAD-FMK was found ineffective in inhibiting SsA-induced cell death in HCC. Importantly, our result indicated that SsA induced the expression of activation transcription factor 3 (ATF3). SsA-induced cell ferroptosis and suppression of SLC7A11 are dependent on ATF3 in HCC. Moreover, we revealed that SsA induced ATF3 upregulation via activation of endoplasmic reticulum (ER) stress. Taken together, our findings support that ATF3-dependent cell ferroptosis mediated the antitumor effects of SsA, opening the possibility to explore SsA as a ferroptosis inducer in HCC.

Circular RNAs as diagnostic biomarkers for gastric cancer: A comprehensive update from emerging functions to clinical significances.

The incidence and mortality of gastric cancer ranks as a fouth leading cause of cancer death worldwide, especially in East Asia. Due to the lack of specific early-stage symptoms, the majority of patients in most developing nations are diagnosed at an advanced stage. Therefore, it is urgent to find more sensitive and reliable biomarkers for gastric cancer screening and diagnosis. Circular RNAs (circRNAs), a novel type of RNAs with covalently closed loops, are becoming a latest hot spot in the field of. In recent years, a great deal of research has demonstrated that abnormal expression of circRNAs was associated with the development of gastric cancer, and suggested that circRNA might serve as a potential biomarker for gastric cancer diagnosis. In this review, we summarize the structural characteristics, formation mechanism and biological function of circRNAs, and elucidate research progress and existing problems in early screening of gastric cancer.

RETRACTED: The role of ATG-7 contributes to pulmonary hypertension by impacting vascular remodeling.

This article has been retracted: please see Elsevier Policy on Article Withdrawal (https://www.elsevier.com/about/our-business/policies/article-withdrawal). At the request of the authors, this article has been retracted following questions of data ownership that were brought to the Editor's attention after publication. Concerns were raised over whether all data were generated in the authors' lab, and over ownership of the models used. The authors were unable to reach a resolution with other labs involved, and as a result all authors have agreed to a retraction of this article.

Keratin 1 attenuates hypoxic pulmonary artery hypertension by suppressing pulmonary artery media smooth muscle expansion.

AIM: Abnormally activated vascular smooth muscle cells are key factors in pulmonary artery remodelling (PAR) and pulmonary artery hypertension (PAH). Keratin 1 is involved in inflammatory diseases; however, its role in PAH is unknown. We speculated that keratin 1 could regulate PASMCs and prevent PAH. METHODS: Rats were exposed to hypoxia (10% O2 ) or MCT (50 mg/kg, intraperitoneal injection) or treated with AAV6 virus. PAR was measured through HE and Masson staining. PASMC activities were measured using MTS assay, EdU and Western blot analyses after cell knockdown with siRNAs or overexpression with Krt1 vectors. RESULTS: 1. Hypoxic PAR was associated with a decrease in keratin 1, especially in PASMCs. 2. Keratin 1 knockdown led to cell proliferation, migration and contraction to synthetic transformation, while keratin 1 overexpression attenuated hypoxia-induced changes in PASMCs. 3. Decreased keratin 1 induced TLR7 upregulation and mediated increases in the inflammatory factors S100a8 and S100a9. 4. Keratin 1 overexpression reduced the inflammatory factor expression induced by TLR7 activation. 5. Further studies demonstrated that keratin 1 expression was negatively correlated with pulmonary vascular pressure following prolonged hypoxia. 6. Pre-treatment with keratin 1 decreased pulmonary artery pressure and the right heart hypertrophy index and alleviated PAR in two model rats. 7. Keratin 1 exhibited a hypermethylation status in hypoxic pulmonary arteries in the sequencing. Hypoxia-induced decrease in keratin 1 expression was associated with Dnmt1 upregulation induced by YY1 downregulation in PASMCs. CONCLUSION: This study suggests that keratin 1 regulates PASMC expansion and has a preventive effect on PAH.

[A case control study of lumbar fusion surgery with the Wiltse approach and the traditional approach].

OBJECTIVE: To compare the clinical results between the Wiltse approach and traditional approach in lumbar fusion. METHODS: The clinical data of 70 patients with lumbar disc herniation or lumbar spondylolisthesis within Meyerding II degree who underwent lumbar fusion surgery from May 2016 to May 2017 were retrospectively analyzed. According to the surgical approach, the patients were divided into Wiltse approach group and traditional approach group. A total of 35 patients in Wiltse approach group, included 18 males and 17 females, with an average age of (52±11) years old;other 35 patients in traditional approach group, included 19 males and 16 females, with an average age of (51±14) years old. Included 38 patients with lumbar disc herniation and 32 patients with spondylolisthesis of Meyerding II degree. The operation time, intraoperative blood loss and postoperative drainage, the VAS score of low back pain and leg pain, the level of creatine phosphokinase (CK) and the cross-sectional area of multifidus muscl on MRI were recorded. RESULTS: The operation time, intraoperative blood loss and postoperative drainage in Wiltse approach group were less than in traditional approach group(P<0.05). There were significant differences in VAS score of low back pain at 7 days and 3 months after operation between two groups(P<0.05). VAS of back pain at both 7 days and 3 months showed better results (P<0.05); VAS of leg pain showed better results in 3 months but had no significant difference in 7 days. There was no significant difference in VAS score of leg pain at 7 days after operation between two groups(P>0.05), but at 3 months had significant difference(P<0.05). The peripheral blood CK levels at 1 day and 3 days after operation respectively were(400±103)U/L and (176±58)U/L in Wiltse approach group, while in traditional approach group were (598±57) U/L and (222±50) U/L, with statistical significance between the two groups(P<0.05). Preoperative cross-sectional area of multifidus muscl on MRI was (424±66) mm² in Wiltse approach group and (428±82) mm² in traditional approach group, there was no significant difference between two groups(P=0.8); at 3 months after operation, in Wiltse approach group was (347±73) mm² and in traditional approach group was(239±78)mm², there was significant difference between two groups(P<0.05). CONCLUSIONS: For lumbar spinal fusion surgery, compared with the traditional approach, Wiltse approach has advantages of shorter operation time, smaller paravertebral muscles injury, and obviously releasing postoperative low back and leg pain. However, in determining the surgery program, the surgical operater also should fully recognize that the anatomical differences of Wiltse approach may influence on operation.

Galectin-3- Mediated Transdifferentiation of Pulmonary Artery Endothelial Cells Contributes to Hypoxic Pulmonary Vascular Remodeling.

BACKGROUND/AIMS: Vascular muscularity is a key event in vessel remodeling during pulmonary artery hypertension (PAH). Endothelial-mesenchymal transdifferentiation (EndMT) has been increasingly reported to play a role in disease occurrence. Galectin-3, a carbohydrate-binding protein regulates cell proliferation, differentiation, migration and neovascularization. However, whether galectin-3 controls endothelial cell transdifferentiation during the development of PAH is unknown. METHODS: Rats were exposed to normoxic or hypoxic conditions (fraction of inspired O2 0.10) for 21 d to establish PAH models. Hemodynamic changes were evaluated through surgery of the right jugular vein and ultrasound biomicroscopy inviVue. And vessel pathological alterations were detected by H&E staining. Galectin-3 (Gal-3)-induced pulmonary artery endothelium cell (PAEC) dynamic alterations were measured by MTT assays, Cell immunofluorescence, Flow cytometry, Real-time PCR and Western blot. RESULTS: Our study demonstrated that Gal-3 was expressed in hypoxic pulmonary vascular adventitia and intima. The increased Gal-3 expression was responsible for hypoxic vessel remodeling and PAH development in vivo. Gal-3 was found to inhibit cell proliferation and apoptosis in cultured endothelial cells. Meanwhile endothelial cell morphology was altered and exhibited smooth muscle-like cell features as demonstrated by the expression of α-SMA after Gal-3 treatment. Gal-3 activated Jagged1/Notch1 pathways and induced MyoD and SRF. When MyoD or SRF were silenced with siRNAs, Gal-3-initiated transdifferentiation in endothelial cells was blocked as indicated by a lack of α-SMA. CONCLUSION: These results suggest that Gal-3 induces PAECs to acquire an α-SMA phenotype via a transdifferentiation process which depends on the activation of Jagged1/Notch1 pathways that mediate MyoD and SRF expression.