南方医科大学学报杂志最新文献

Objectives: To investigate the role of Cav1 gene in regulating mitochondrial function and mRNA m⁶A modification and expressions of the key genes in mouse hepatocytes.

Methods: In mouse hepatocyte AML12 cells, the effects of lentivirus-mediated Cav1 knockdown and transfection with a negative control virus on mitochondrial membrane potential and mitochondrial respiratory function were analyzed using the TMRE fluorescent probe and the Seahorse extracellular flux analysis system. Methylated RNA immunoprecipitation (MeRIP) combined with m⁶A microarray was utilized to evaluate the changes in mRNA m⁶A modification and expression levels, followed by enrichment analysis to identify the functionally relevant genes. The m⁶A modification and expression levels of the mRNAs were validated by qPCR.

Results: Compared with the negative control group, the cells with Cav1 knockdown exhibited significantly reduced mitochondrial membrane potential and respiratory capacity. m⁶A microarray analysis revealed significant changes in m⁶A modification levels (fold change>1.5) of 7814 mRNAs, including 152 upregulated and 7662 downregulated mRNAs. Integrated expression analysis identified 2497 mRNAs showing coordinated changes in m⁶A modification and expression levels. These mRNAs were enriched in the oxidative phosphorylation pathway, with Usp15, Suclg2, and Ppa2 exhibiting the highest percent changes in m⁶A modification. Both microarray and qPCR results showed that the m⁶A modification and expression levels of Usp15, Suclg2 and Ppa2 mRNAs were significantly reduced in cells with Cav1 knockdown compared to the NC group.

Conclusions: Cav1 knockdown induces significant alterations in mRNA m⁶A modification as well as their expression levels. The regulatory effects of Cav1 on mitochondrial function may be mediated by modulation of m⁶A modification of Usp15, Suclg2, and Ppa2 mRNAs.

Objectives: To investigate the clinical significance of abnormal expression of angio-associated migratory cell protein (AAMP) in hepatocellular carcinoma (HCC).

Methods: Bioinformatics analyses were used to analyze AAMP expression level in HCC and its prognosis value. In 60 pairs of HCC and adjacent tissues, AAMP expression was detected immunohistochemically and its correlation with clinicopathological characteristics of the patients was analyzed. In cultured Mahlavu and Huh-7 cells with lentivirus-mediated AAMP knockdown, the changes in cell proliferation, apoptosis, migration and invasion were observed, and their lung metastasis following tail vein injection in nude mice were assessed. In HCC cells with AAMP knockdown, Western blotting, immunofluorescence staining or RT-qPCR were used to examine the changes in expression levels of E-cadherin, N-cadherin, Vimentin and Snail and the effects of MG-132 and CHX on RhoA expression. The correlation between the expressions of AAMP and RhoA in HCC tissues was analyzed by immunohistochemistry.

Results: Bioinformatics analysis showed that AAMP expression was elevated in HCC tissues (P<0.05) in correlation with advanced clinical stage and poor prognosis (P<0.05). Immunohistochemistry results confirmed significant correlations of high AAMP expression with Edmondson-Steiner grade (III+IV), venous infiltration and TNM stage (III+IV) of HCC (P<0.05). In cultured HCC cells, AAMP knockdown did not significantly affect cell proliferation or apoptosis, but obviously suppressed cell migration and invasion in vitro and lung metastasis in nude mice. AAMP knockdown significantly increased E-cadherin expression, decreased N-cadherin, Vimentin and Snail expressions, and reduced RhoA protein levels without obviously affecting RhoA mRNA levels. MG-132 treatment blocked the inhibitory effect of AAMP knockdown on RhoA protein expression. The expressions of AAMP and RhoA showed a significant positive correlation in HCC tissues (P<0.05).

Conclusions: AAMP overexpression is associated with malignant clinical features of HCC and promotes epithelial-mesenchymal transition and metastasis of HCC cells partly by stabilizing RhoA expression.

Objectives: To explore the mechanism of Shenqi Xiezhuo Decoction (SQXZD) for improving renal fibrosis (RF) in rats.

Methods: The chemical components of SQXZD were identified using UPLC-Q Exactive/MS, and component-disease target network and enrichment analyses were conducted to screen the key pathways and targets. In the animal experiment, 49 male SD rats were randomized equally into blank control group, sham operation group, unilateral ureteral obstruction-induced RF model group, losartan treatment (daily dose 4.6 mg/kg) group, and low-, medium-, and high-dose SQXZD (9.7, 19.4, and 38.8 g/kg, respectively) treatment groups. After 14 days' treatment, renal pathologies and collagen deposition of the rats were examined with HE and Masson staining, and serum levels of BUN, Cr, SOD, MDA, GSH-px, IL-6, and TNF-α were detected. Western blotting and qRT-PCR were used to detect renal protein and mRNA expressions of α‑SMA, Col-I, NAKED2, Rap1, B-raf, Raf-1, MEK3/6, p38MAPK, MEK, ERK1/2, p-ERK1/2, FoxO3a, p-FoxO3a, and MnSOD.

Results: A total of 263 chemical components were identified in SQXZD. Network pharmacology revealed 170 intersecting targets between the components and RF enriched in the MAPK, Rap1, and FoxO pathways. The rat models of RF showed abnormal renal structural changes, increased fibrosis area, elevated serum BUN, Cr, MDA, IL-6, and TNF-α levels, reduced SOD and GSH-px levels, upregulated renal expressions of α‑SMA, Col-I, NAKED2, Rap1, B-raf, MEK, ERK1/2, p-ERK1/2, MEK3/6, and p38MAPK, and downregulated Raf-1, FoxO3a, p-FoxO3a, and MnSOD expressions. Treatment with losartan and SQXZD (especially at the medium dose) obviously lessened renal pathologies, improved renal functions, alleviated oxidative stress and inflammation, and ameliorated abnormal changes in the Rap1/MAPK/FoxO3a signaling pathway in the rat models.

Conclusions: SQXZD alleviates RF and improves renal function in rats possibly by ameliorating renal oxidative stress and inflammation via regulating the Rap1/MAPK/FoxO3a signaling pathway.

Objectives: To investigate the inhibitory effect of polyphyllin VII (PP7) on osteosarcoma xenograft growth in mice and explore the underlying molecular mechanism.

Methods: Ultra‑performance liquid chromatography‑tandem mass spectrometry was used to analyze the main active components of Paris polyphylla. Six nude mice bearing patient‑derived xenograft (PDX) were randomized into two groups for treatment with 2 mg/kg PP7 gavage or saline every other day for 28 days, and the changes in tumor volume and mass were measured. In cultured 143B and HOS cells, the effect of PP7 treatment (0, 1.25, 2.5, 5, and 10 μmol/L) on cell proliferation was assessed with CCK‑8 assay, and Transwell assays were employed to examine the changes in cell migration and invasion. The target of PP7 was predicted by integrated analyses with single‑cell RNA sequencing (scRNA‑seq), bulk RNA sequencing (bulk RNA‑seq) and molecular docking and verified using Western blotting. In osteosarcoma cells transfected with SOHLH1 siRNAs or a negative control sequence, the effects of PP7 treatment (5 μmol/L) on cell migration, invasion, ferroptosis, reactive oxygen species (ROS) production and lipid peroxidation (LPO) were analyzed.

Results: PP7 was identified as one of the major active constituents of Paris polyphylla. In the tumor-bearing mice, PP7 treatment significantly lower the tumor volume and mass. In 143B and HOS cells, PP7 concentration‑dependently inhibited cell proliferation, and at 5 μmol/L, PP7 significantly inhibited cell proliferation, migration and invasion. Multi‑omics analysis identified SOHLH1 as a potential target of PP7, and Western blotting confirmed that PP7 upregulated SOHLH1 expressions at both the mRNA and protein levels. SOHLH1 silencing obviously attenuated the inhibitory effects of PP7 on cell migration and invasion and reduced PP7‑induced ferroptosis.

Conclusions: PP7 suppresses osteosarcoma xenograft growth in mice by inducing ferroptosis via upregulating SOHLH1 expression.

Objectives: To investigate the mechanism by which Morinda officinalis polysaccharide (MOP) delays osteoarthritis chondrocyte degeneration.

Methods: In primary cultures of chondrocytes from 4-week-old C57BL/6 mice, the effects of IL-1β and MOP treatment at different concentrations on cell viability were assessed with CCK-8 assay. The treated cells were examined for protein expressions of PKM2, caspase-1, and GSDMD using Western blotting and for XIST expression using fluorescence in situ hybridization (FISH). In IL-1β-induced mouse chondrocytes, the effects of MOP, transfection for XIST overexpression or knockdown, and MOP treatment after the transfection were tested by detecting mRNA levels of GluT1, HK2, PKM2, LDHA, PFKFB3, NLRP3, caspase-1, and GSDMD; flow cytometry, Western blotting, and toluidine blue staining were used to analyze chondrocyte apoptosis, expressions of glycolysis and pyroptosis regulators, and glycosaminoglycan expression.

Results: The second-passage chondrocytes showed good viability and positive collagen II staining. IL-1β induction caused degenerative morphological changes of the cells, decreased collagen II expression, and upregulated cellular expressions of PKM2, caspase-1, and GSDMD proteins. MOP treatment (especially at 4 mg/mL) significantly enhanced cell viability and reduced HK2, PKM2, caspase-1 and GSDMD expressions in IL-1β‑induced mouse chondrocytes. XIST was localized predominantly in the nuclei of the chondrocytes, and its expression increased significantly in IL-1β‑treated cells, and was attenuated by MOP treatment. XIST overexpression synergized with IL-1β to upregulate mRNA and protein expressions of glycolysis- and pyroptosis-related factors in the chondrocytes, and such effects were obviously attenuated by MOP. Conversely, XIST knockdown significantly inhibited chondrocyte apoptosis and glycosaminoglycan expression, and down-regulated glycolysis- and pyroptosis-related proteins. MOP treatment exhibited similar protective effects to XIST knockdown, and their combination significantly augmented these protective effects.

Conclusions: MOP mitigates IL-1β‑induced mouse chondrocyte degeneration by modulating glycolysis and pyroptosis via targeting XIST.

Objectives: To investigate the regulatory effect of Serpin Family E Member 1 (SERPINE1) on immune microenvironment and paclitaxel (PTX) resistance of triple-negative breast cancer (TNBC) cells.

Methods: CCK-8 assay was used to determine the half-maximal inhibitory concentration of PTX in TNBC cell line MDA-MB-231. In wild-type MDA-MB-231 cells and a PTX-resistant MDA-MB-231 cell line (MDA-MB-231/PTX) established by stepwise increasing low-dose PTX treatment, the effects of Western blot-verified transfection with SERPINE1 overexpression plasmids or SERPINE1 siRNAs on cell apoptosis were evaluated using Hoechst 33258 staining and by detecting expression levels of cleaved caspase-3 using Western blotting. The changes in proliferation of the transfected cells were assessed using EdU and CCK-8 assays. The breast cancer cells with different treatments were co-cultured with macrophages, and M1 and M2 polarization of the macrophages were analyzed with flow cytometry and Western blotting. In nude mouse models bearing subcutaneous breast cancer cell xenografts, the effects of SERPINE1 overexpression and knockdown in the engrafted cells on tumor growth and PTX resistance were evaluated.

Results: SERPINE1 overexpression significantly inhibited apoptosis and promoted proliferation of MDA-MB-231 cells, and SERPINE1 knockdown obviously promoted apoptosis and inhibited proliferation of MDA-MB-231/PTX cells. The macrophages co-cultured with SERPINE1-overexpressing breast cancer cells showed enhanced M2 polarization and suppressed M1 polarization with a lowered M1/M2 ratio. In the tumor-bearing nude mouse models, SERPINE1 overexpression in the engrafted cells resulted in significantly accelerated tumor growth.

Conclusions: In MDA-MB-231 cells, SERPINE1 overexpression promotes cell proliferation, inhibits apoptosis, and enhances PTX resistance. SERPINE1 plays a regulatory role in macrophage polarization in the immune microenvironment of breast cancer, and its high expression promotes M2 polarization of the macrophages.

Objectives: To evaluate the effect of resveratrol (RES) on barrier function of mouse brain microvascular endothelial cell monolayers exposed to oxygen/glucose deprivation/reoxygenation (OGD/R) and PM_2.5 and explore the role of mitochondrial fission and fusion in protecting endothelial barrier function.

Methods: Cultured mouse brain microvascular endothelial cells were exposed to OGD/R, treated with PM_2.5 (100 μg/mL) before OGD/R, or pretreated with RES (40 mg/mL) prior to OGD/R+PM_2.5 exposures. The changes in cell viability were examined with CCK-8 assay, and cell permeability was assessed by measuring transendothelial electrical resistance (TEER) and FITC-dextran permeation. Malondialdehyde (MDA) content and superoxide dismutase (SOD) activity were measured, and intracellular and mitochondrial ROS levels were detected using fluorescent probes. Mitochondrial morphology in the treated cells was observed using Mito-Tracker Red CMXRos. Western blotting was performed to detect the changes in cellular expressions of the tight junction proteins (ZO-1, occludin, and claudin-5) and mitochondrial dynamics-associated proteins (Drp1, Fis1, Mfn2, and OPA1).

Results: Compared with the normal control cells, the cells exposed to OGD/R or both OGD/R and PM_2.5 showed significantly decreased TEER levels, increased permeability, elevated oxidative stress, and increased ROS fluorescence intensities. Obvious mitochondrial fragmentation and morphological changes in the mitochondria were observed in the exposed cells, which also showed decreased expressions of tight junction proteins and mitochondrial fusion proteins with increased expressions of mitochondrial fission proteins. RES pretreatment of the endothelial cells before the exposures significantly reduced membrane permeability, lowered ROS levels, improved mitochondrial morphology, increased expressions of tight junction and fusion proteins, and decreased fission protein expressions.

Conclusions: RES can protect barrier function of mouse brain microvascular endothelial cell monolayers exposed to OGD/R and PM_2.5 by modulating mitochondrial dynamics, potentially through promoting mitochondrial fusion and inhibiting mitochondrial fission.

Objectives: To investigate the causal relationship between gut microbiota, T-cell function, and the risk of colorectal cancer.

Methods: Gut microbiota data from the MiBioGen database and T-cell and colorectal cancer data from publicly available GWAS datasets were obtained for analyzing the causality between gut microbiota, T-cell subsets, and the risk of colorectal cancer with two-sample Mendelian randomization (MR) analyses, using inverse variance weighting as the primary analytical method supplemented with MR-Egger, weighted median, simple mode, and weighted mode methods. Horizontal pleiotropy was assessed using MR-PRESSO and MR-Egger regression. Cochran's Q test was used to evaluate heterogeneity, and sensitivity analysis was performed using the leave-one-out method.

Results: In the Forward MR analysis of gut microbiota and T cells, 11 gut microbiota species showed causal relationships. Six of these species exhibited positive correlations with T cells, including Prevotella7 (P=0.003), Ruminococcaceae UCG011 (P=0.033), Ruminococcaceae UCG004 (0.010), Ebacterium brachy group (P=0.005), Lachnospiraceae FCS020 group (P=0.028), and Coprobacter (P=0.033), and the remaining 5 species showed negative correlations with T cells. Forward MR analysis of T cells and colorectal cancer suggested that CD25⁺⁺CD45RA^-CD4⁺ non-regulatory T cells were negatively correlated with colorectal cancer risk (IVW: OR=0.935, 95% CI: 0.878-0.995; P=0.035). The analysis of gut microbiota and colorectal cancer suggested that 11 gut microbiota species were causally associated with colorectal cancer, and 6 of them (Eubacterium xylanophilum group, P=0.039; Selenomonadales, P=0.014; Negativicutes, P=0.014; Bifidobacteriaceae, P=0.048; Bifidobacteriales, P=0.048; and Coprococcus1, P=0.033) showed positive correlations and the remaining 5 showed negative correlations.

Conclusions: Coprobacter spp. and Eubacterium xylanophilum group spp. are causally associated with both T cell activity and colorectal cancer risk, and the former bacteria induce inactivation of CD25⁺⁺CD45RA^-CD4⁺ non-regulatory T cells to promote colorectal cancer progression, whereas the latter bacteria promote CD25⁺⁺CD45RA^-CD4⁺ non-regulatory T cell activity to inhibit colorectal cancer development.

Objectives: To investigate the potential mechanism underlying the protective effect of secoisolariciresinol diglucoside (SDG) against chronic kidney disease (CKD) in offspring mice caused by maternal exposure to trans fatty acids (TFA) during pregnancy and lactation.

Methods: Thirty female C57BL/6 mice were randomized into control group, TFA model group, and 3 TFA model groups treated with SDG at low, medium and high doses (10, 20 and 30 mg/kg, respectively). The changes in blood urea nitrogen (BUN) and serum creatinine (CRE) levels of the mice were measured. Network pharmacology analysis was conducted to explore protective mechanism of SDG against TFA-induced renal injury, and molecular docking was used to assess the binding affinity of SDG to Bcl-2, Bax, and caspase-3. The protein expressions of cleaved caspase-3, Bax, and Bcl-2 in the renal tissues of the offspring mice were detected with Western blotting.

Results: The mice in TFA group showed significantly higher BUN and CRE levels than those in the control group. Treatment with SDG at the medium and high doses significantly reduced BUN and CRE levels in the mouse models. Network pharmacology and molecular docking suggested that SDG ameliorated renal injury by targeting the apoptosis-related Bcl-2/Bax/caspase-3 axis. The results of Western blotting showed the mouse models in TFA exposure group had increased renal cell apoptosis with elevated expression levels of cleaved caspase-3 protein and a decreased Bcl-2/Bax ratio (P<0.05), and intervention with SDG at all the 3 doses significantly reduced renal cell apoptosis and renal expression of cleaved caspase-3 and increased the Bcl-2/Bax ratio in the mouse models.

Conclusions: Maternal TFA exposure during gestation and lactation induces renal injury in offspring mice. Dietary SDG intervention can mitigate TFA-induced renal injury in offspring mice possibly by suppressing renal cell apoptosis via regulating the Bcl-2/Bax/caspase-3 signaling axis.

Objectives: To investigate YEATS2 expression in gastric cancer (GC), its prognostic value, and its regulatory role in epithelial-mesenchymal transition (EMT) of GC cells.

Methods: YEATS2 expression in GC was analyzed using publicly available databases. Paired GC and adjacent tissues were collected from 100 patients undergoing radical surgery for immunohistochemical detection of YEATS2 expression, and its correlations with the patients' clinicopathological parameters and Ki67 expression were analyzed. The prognostic value of YEATS2 was assessed using Kaplan-Meier analysis, Cox regression and ROC curves, and its regulatory mechanisms were analyzed using KEGG enrichment analysis. In cultured GC cell lines (HGC-27 and AGS), the effect of YEATS2 knockdown and overexpression on migration, invasion and EMT of the cells were examined with scratching assay, Transwell assay and Western blotting.

Results: YEATS2 was significantly overexpressed in GC tissues with a positive correlation with Ki67 (P<0.05). High YEATS2 expression was associated with elevated CEA (≥5 μg/L), CA19-9 (≥37 kU/L), T3-4 stage, and N2-3 stage (all P<0.05). Patients with high YEATS2 expression had significantly reduced 5-year survival (P<0.001); ROC analysis showed that YEATS2 expression levels had a sensitivity of 80.00% and a specificity of 66.67% for predicting patient survival (P<0.05). Cox regression identified high YEATS2 as an independent risk factor for poor postoperative 5-year survival outcome of GC patients (HR: 1.675, 95%CI: 1.013-2.771; P=0.045). KEGG enrichment analysis suggested involvement of YEATS2 in EMT in GC and Wnt/β-catenin signaling. In cultured GC cells, YEATS2 overexpression significantly promoted cell migration and invasion, upregulated the expressions of vimentin, N-cadherin, Wnt and active β-catenin, and downregulated E-cadherin expression, and these changes were obviously suppressed by treatment with XAV-939 (a Wnt/β-catenin inhibitor).

Conclusions: High YEATS2 expression activates Wnt/β-catenin signaling to promote EMT in GC and is correlated with poor prognosis of GC patients.