Folia histochemica et cytobiologica最新文献

Introduction: Prognostic and predictive value of PD-L1 as a biomarker in breast cancer remains controversial. While some studies suggest its association with negative prognostic parameters, others reported a highly significant association between PD-L1 expression and tumor-infiltrating lymphocytes, which are known to be an independent favorable prognostic factor. The aim of present study is to examine the relationship between immune response markers and PD-L1 expression in early breast cancer.

Material and methods: Immunohistochemical expression of PD-L1, along with density and composition of stromal lymphocytic infiltrate and peritumoral lymphoid aggregates was analyzed in 95 samples of invasive breast cancer.

Results: A strong positive correlation between PD-L1 expression and the density of stromal lymphocytic infiltrate and peritumoral lymphoid aggregates was identified and a cut-off value of 53% coverage of tumor stroma by lymphocytes, with which PD-L1 positivity can be predicted with excellent diagnostic accuracy, was determined for the first time using statistical methods. Additionally, PD-L1 positivity was observed significantly more often in tumors with higher absolute number of both CD4 and CD8 T-lymphocytes in the stromal infiltrate. No significant correlation with molecular subtype of breast cancer was found.

Conclusions: Our results indicate that the density of stromal lymphocytic infiltrate might be a better predictor of PD-L1 positivity in early breast cancer than the molecular subtype and that the key to the optimization of PD-L1 as a biomarker in breast cancer lies in its interpretation in the context of other immune response markers.

Introduction: Transplantation of mesenchymal stem cells (MSCs) has been reported to be a novel promising target for the regeneration of degenerated intervertebral discs (IVDs). However, the culture and survival limitations of MSCs remain challenging for MSC-based biological therapy. Myricetin, a common natural flavonoid, has been suggested to possess antiaging and antioxidant abilities. Therefore, we investigated the biological function of myricetin, and its related mechanisms involving cell senescence in intervertebral disc degeneration (IDD).

Material and methods: The nucleus pulposus-derived mesenchymal stem cells (NPMSCs) were isolated from 4-month-old Sprague-Dawley (SD) rats and identified by examining surface markers and multipotent differentiation. Rat NPMSCs were cultured in an MSC culture medium or culture medium with different concentrations of H2O2. Myricetin or the combination of myricetin and EX527 were added to the culture medium to investigate the effects of myricetin. Cell viability was evaluated by cell counting kit-8 assays (CCK-8). The apoptosis rate was determined using Annexin V/PI dual staining. The mitochondrial membrane potential (MMP) was analyzed by a fluorescence microscope after JC-1 staining. The cell senescence was determined by SA-β-Gal staining. MitoSOX green was used to selectively estimate mitochondrial reactive oxygen species (ROS) Apoptosis-associated proteins (Bax, Bcl2, and cleaved caspase-3), senescence markers (p16, p21, and p53), and SIRT1/PGC-1α signaling pathway-related proteins (SIRT1 and PGC-1α) were evaluated by western blotting.

Results: The cells isolated from nucleus pulposus (NP) tissues met the criteria for MSCs. Myricetin showed no cytotoxicity up to a concentration of 100 μM in rat NPMSCs cultured for 24 h. Myricetin pretreatment exhibited protective effects against H₂O₂-induced apoptosis. Myricetin could also alleviate H₂O₂-induced mitochondrial dysfunctions of increased mitochondrial ROS production and reduced MMP. Moreover, myricetin pretreatment delayed rat NPMSC senescence, as evidenced by decreased exppression of senescence indicators. Pretreatment of NPMSCs with 10 μM EX527, a selective inhibitor of SIRT1, prior to exposure to 100 μM H2O2, reversed the inhibitory effects of myricetin on cell apoptosis.

Conclusions: Myricetin could affect the SIRT1/PGC-1α pathway to protect mitochondrial functions and alleviate cell senescence in H₂O₂-treated NPMSCs.

Adipose mesenchymal stem cell-derived exosomes (ADMSC-Exo) are a new strategy for the treatment of liver injury. However, mesenchymal stem cells (MSCs) exert therapeutic effects mainly by secreting hepatocyte growth factor (HGF). Therefore, we investigated the role of exosomes derived from ADMSC that overexpress HGF (ADMSCHGF-Exo) on liver injury.

Material and methods: ADMSCs were isolated from young BALB/c female mice. Then exosomes derived from ADMSC transfecting negative control (ADMSCNC-Exo) and HGF overexpression (ADMSCHGF-Exo) were isolated and identified by quantitative polymerase chain reaction (qPCR), flow cytometry, western blot, transmission electron microscope and Nanosight particle tracking analysis. These exosomes were injected into male mice via tail vein after inducing liver injury by administering 40% carbon tetrachloride (CCl₄)-olive oil twice a week (3 mL/kg, subcutaneously) for 6 weeks. Liver injury and liver collagen fiber accumulation were determined by histopathological analysis. Then, the levels of serum liver function indexes (alanine aminotransferase, aspartate aminotransferase, albumin, total bilirubin), hepatocyte-specific markers (albumin, cytokeratin-18 and hepatocyte nuclear factor 4α), hepatic fibrosis-related proteins (α-smooth muscle actin and collagen I) and Rho GTPase (cell division cycle 42 and ras-related C3 botulinum toxin substrate 1) were determined by Enzyme-linked immunosorbent assay (ELISA), immunohistochemistry, Western blot and qPCR.

Results: ADMSCs were identified by high expression of CD105 and CD44 molecules and low expression of CD45 and CD34. ADMSCs-Exo, ADMSCNC-Exo and ADMSCHGF-Exo transfected cells had similar expression of exosome-specific membrane proteins (CD63, CD81 and CD9). Mice with CCl₄-induced liver injury exhibited abnormal serum liver function indexes, altered expression of hepatocyte-specific markers, hepatic fibrosis-related proteins and Rho GTPase protein as well as histopathological changes and collagen fiber accumulation in the liver. These changes were reversed by ADMSC-Exo, ADMSCNC-Exo and ADMSCHGF-Exo administration with ADMSCHGF-Exo displaying the most significant impact.

Conclusions: ADMSCHGF-Exo exerted a hepatoprotective effect in mice with experimental liver injury by alleviating hepatic fibrosis and restoring liver function.

Introduction: Sonodynamic therapy (SDT), a promising non-invasive therapeutic modality, has attracted increasing attention in the treatment of pancreatic cancer (PC). At present, the role of autophagy in SDT of PC remains unclear. This study aims to explore the role of autophagy in SDT of PC and its effect on apoptosis of PC cells.

Material and methods: PC cells (Capan-1 and BxPC-3) underwent incubation with 5-aminolevulinic acid (5-ALA) or/and ultrasound (US) exposure (control, 5-ALA, US, and SDT groups), followed by measurement of cell apoptosis and autophagy. Specifically, cell viability, apoptosis, and the expression of apoptosis-related proteins (cleaved Caspase-3, Bax, and Bcl-2) were measured using CCK-8 assay, flow cytometry, and western blot analysis, respectively. The mitochondrial morphology was observed with the transmission electron microscopy, accompanied by the detection of autophagosome marker (LC3) co-located with Mito and the protein expression of LC3II/I. Before SDT treatment, the autophagy inhibitor 3-MA and the apoptosis inhibitor z-VAD were respectively added to PC cell cultures to evaluate the effects of autophagy inhibition on apoptosis and apoptosis inhibition on autophagy in PC cells.

Results: Compared with the control group, cell viability was inhibited and cell apoptosis and autophagy were enhanced in the SDT group, while cell viability, autophagy, and apoptosis in the 5-ALA and US groups were not significantly changed. Moreover, 3-MA treatment inhibited autophagy and accelerated apoptosis, whereas z-VAD treatment reduced apoptosis but did not affect autophagy in PC cells.

Conclusions: Autophagy was activated in SDT-treated PC cells, and inhibition of autophagy promoted cell apoptosis in PC cells.

Introduction: High-fructose, high-fat diet consumption (HFHF) is one of the primary causes of non-alcoholic fatt liver disease (NAFLD), which is due to impaired beta-oxidation or apolipoprotein secretion by hepatocytes. Activation of autophagy in hepatocytes could be a therapeutic method against hepatic complications. This study was designed to compare effects of rapamycin and intermittent fasting-inducing autophagy in rats with experimentally induced nonalcoholic fatty liver.

Material and methods: Male rats were divided into five groups: C (control, n = 6), the experimental group (EX) subdivided, EXIa (HFHF, n = 6), EXIb (recovery, n = 6), EXII (rapamycin, n = 6) and EXIII (intermittent fasting, n = 6). All rats in the experimental group received HFHF diet for 8 weeks to induce nonalcoholic-fatty liver and obesity. Then, for the next 8 weeks the animals received either a daily oral dose of rapamycin (EXII group) or to intermittent fasting (IF) for 16 hours daily (EXIII group). Blood samples were drawn, and serum TG concentration as well as ALT and AST activities were determined. Hepatic sections were examined by light and electron microscopy. LC3B immunohistochemical staining, morphometric and statistical studies were performed.

Results: Subgroups EXIa (HFHF subgroup) and EXIb (Recovery subgroup) showed marked increase in TG, ALT, and AST levels associated with loss of normal hepatic architecture, cytoplasmic vacuolations and faint LC3B immunoreactivity. Ultrathin sections exhibited many autophagosomes in hepatocytes. On the other hand, rapamycin (EXII) and IF (EXIII) groups showed significant improvement to a variable extent in comparison to EXI groups.

Conclusions: It could be concluded that rapamycin and intermittent fasting significantly improved NAFLD-induced changes of liver structure and function by inducing autophagy in hepatocytes.