Experimental Biology and Medicine

Vitamin D₃ has been reported to protect liver against non-alcoholic fatty liver disease (NAFLD) by attenuating hepatic lipid dysregulation in type 2 diabetes mellitus (T2DM). However, the mechanism of vitamin D₃ on hepatic lipid metabolism-associated autophagy in hyperglycemia-induced NAFLD remains yet to be exactly elucidated. C57BL/6J mice were intraperitoneally injected with 30 mg/kg of streptozotocin and fed a high-fat diet for induction of diabetes. All mice were administered with vehicle or vitamin D₃ (300 ng/kg or 600 ng/kg) by oral gavage for 12 weeks. Histological demonstrations of the hepatic tissues were obtained by H&E staining and the protein levels related to lipid metabolism and autophagy signaling were analyzed by Western blot. Treatment with vitamin D₃ improved insulin resistance, liver damage, and plasma lipid profiles, and decreased hepatic lipid content in the diabetic mice. Moreover, vitamin D₃ administration ameliorated hepatic lipid dysregulation by downregulating lipogenesis and upregulating lipid oxidation under diabetic condition. Importantly, vitamin D₃ treatment induced autophagy by activating AMP-activated protein kinase (AMPK), inactivating Akt and ultimately blocking mammalian target of rapamycin (mTOR) activation in the T2DM mice. Additionally, vitamin D₃ was found to be effective in anti-apoptosis and anti-fibrosis in the liver of diabetic mice. The results suggested that vitamin D₃ may ameliorate hepatic lipid dysregulation by activating autophagy regulatory AMPK/Akt-mTOR signaling in T2DM, providing insights into its beneficial effects on NAFLD in type 2 diabetic patients.

This study aimed to evaluate the role of APOE polymorphisms (rs429358 and rs7412) in the risk of age-related macular degeneration in a sample of the Southeastern Brazilian population. Seven hundred and five unrelated individuals were analyzed, 334 with age-related macular degeneration (case group), and 371 without the disease (control group). In the case group, patients were further stratified according to disease phenotypes, divided into dry and wet age-related macular degeneration, and non-advanced and advanced age-related macular degeneration. APOE polymorphisms (rs429358 and rs7412) were evaluated through polymerase chain reaction and direct sequencing. In the comparison of cases vs. controls, none of the associations reached statistical significance, considering the Bonferroni-adjusted P-value, although there was a suggestive protection for the E3/E4 genotype (OR = 0.626; P-value = 0.037) and E4 carriers (OR = 0.6515; P-value = 0.047). Statistically significant protection for both the E3/E4 genotype and E4 carriers was observed in the comparisons: advanced age-related macular degeneration vs. controls (OR = 0.3665, P-value = 0.491 × 10⁻³ and OR = 0.4031, P-value = 0.814 × 10⁻³, respectively), advanced age-related macular degeneration vs. non-advanced age-related macular degeneration (OR = 0.2529, P-value = 0.659 × 10⁻⁴ and OR = 0.2692, P-value = 0.631 × 10⁻⁴, respectively). In the comparison of wet age-related macular degeneration vs. control, protection was statistically significant only for E3/E4 (OR = 0.4052, P-value = 0.001). None of the comparisons demonstrated any significant association for E2 genotypes or E2 carriers in age-related macular degeneration risk in this study. Findings suggest a protective role of the E4 haplotype in the APOE gene in the risk for advanced and wet forms of age-related macular degeneration, in a sample of the Brazilian population. To our knowledge, this is the first Brazilian study to show the association between APOE polymorphisms and age-related macular degeneration.

Cancer-associated sarcopenia is a complex metabolic syndrome marked by muscle mass wasting. Muscle wasting is a serious complication that is a primary contributor to cancer-related mortality. The underlying molecular mechanisms of cancer-associated sarcopenia have not been completely described to date. In general, evidence shows that the main pathophysiological alterations in sarcopenia are associated with the degradation of cellular components, an exceptional inflammatory secretome and mitochondrial dysfunction. Importantly, we highlight the prospect that several miRNAs carried by tumor-derived exosomes that have shown the ability to promote inflammatory secretion, activate catabolism, and even participate in the regulation of cellular degradation pathways can be delivered to and exert effects on muscle cells. In this review, we aim to describe the current knowledge about the functions of exosomal miRNAs in the induction of cancer-associated muscle wasting and propose potential treatment strategies.

Recurrent epithelial erosion and refractory corneal ulcer are the clinical features of diabetic keratopathy (DK), which eventually lead to corneal scar and visual disturbance. In this study, we sought to determine the abnormalities of cell junction in diabetic corneal epithelial cells and the effect of high glucose on the β-catenin/E-cadherin complex. Corneal histology showed that corneal epithelial cells of high glucose mice were loosely arranged, and the immunohistochemistry showed that the expression of E-cadherin decreased, the levels of β-catenin increased in nuclear. High glucose-induced degradation and endocytosis of E-cadherin of corneal epithelial cells reduce the formation of β-catenin/E-cadherin complex and promote the nuclear translocation of β-catenin. Moreover, high glucose also activated the transcription and expression of matrix metallopeptidase and snail, which interfered with the adhesion of corneal epithelial cells to the basement membrane. These findings reveal that DK is associated with the dissociation of cell junctions. The maintenance of the stability of the β-catenin/E-cadherin complex may be a potential therapeutic target of refractory corneal ulcers in patients with diabetes.

Osteoblasts play an important role in bone regeneration and repair. The hypoxia condition in bone occurs when bone undergoes fracture, and this will trigger a series of biochemical and mechanical changes to enable bone repair. Hence, it is interesting to observe the metabolites and metabolism changes when osteoblasts are exposed to hypoxic condition. This study has looked into the response of human osteoblast hFOB 1.19 under normoxic and hypoxic conditions by observing the cell growth and utilization of metabolites via Phenotype MicroArrays™ under these two different oxygen concentrations. The cell growth of hFOB 1.19 under hypoxic condition showed better growth compared to hFOB 1.19 under normal condition. In this study, osteoblast used glycolysis as the main pathway to produce energy as hFOB 1.19 in both hypoxic and normoxic conditions showed cell growth in well containing dextrin, glycogen, maltotriose, D-maltose, D-glucose-6-phospate, D-glucose, D-mannose, D-Turanose, D-fructose-6-phosphate, D-galactose, uridine, adenosine, inosine and α-keto-glutaric acid. In hypoxia, the cells have utilized additional metabolites such as α-D-glucose-1-phosphate and D-fructose, indicating possible activation of glycogen synthesis and glycogenolysis to metabolize α-D-glucose-1-phosphate. Meanwhile, during normoxia, D-L-α-glycerol phosphate was used, and this implies that the osteoblast may use glycerol-3-phosphate shuttle and oxidative phosphorylation to metabolize glycerol-3-phosphate.

Roundabout guidance receptor proteins are crucial components of the SLIT/ROBO signaling pathway. This pathway is important for the nervous system and in embryonic development. Recently, increasing evidence has shown that roundabout guidance receptor proteins and the SLIT/ROBO signaling pathway also participate in tumorigenesis. Here, by analyzing transcriptome data from the TCGA and GEO databases, we found that ROBO3 is highly expressed in non-M3 acute myeloid leukemia. High ROBO3 expression was associated with increased age at diagnosis and poorer risk classification (both P < 0.01). Patients with high ROBO3 expression had higher rates of TP53 and RUNX1 mutations (both P < 0.05). Significantly worse overall survival and event-free survival were observed in high ROBO3 expression patients compared with low ROBO3 expression patients (OS: P = 0.004; EFS: P= 0.012). High ROBO3 expression was also associated with poorer overall survival and event-free survival in a subgroup of patients who received intensive chemotherapy (OS: P = 0.024; EFS: P = 0.040). Moreover, multivariate analysis indicated that high ROBO3 expression was an independent risk factor for poor overall survival in non-M3 acute myeloid leukemia patients who are younger than 60 and received intensive chemotherapy during remission induction. Bioinformatics analysis by Kyoto Encyclopedia of Genes and Genomes and Gene Ontology revealed that high ROBO3 expression significantly altered cell adhesion and extracellular matrix-related pathways (adjusted P < 0.05). Taken together, the data demonstrate that ROBO3 is upregulated in non-M3 acute myeloid leukemia and may be a potent biomarker of inferior prognosis.

White matter lesion (WML) is caused by chronic cerebral hypoperfusion, which are usually associated with cognitive impairment. Evidence from recent studies has shown that ginkgolide B has a neuroprotective effect that could be beneficial for the treatment of ischemia; however, it is not clear whether ginkgolide B has a protective effect on WML. Our data show that ginkgolide B can promote the differentiation of oligodendrocyte precursor cell (OPC) into oligodendrocytes and promote oligodendrocyte survival following a WML. Ginkgolide B (5, 10, 20 mg/kg) or saline is administered intraperitoneally every day after WML. After 4 weeks, the data of Morris water maze suggested that rats’ memory and learning abilities were impaired, and the administration of ginkgolide B enhanced behavioral achievement. Also, treatment with ginkgolide B significantly attenuated this loss of myelin. Our result suggests that ginkgolide B promotes the differentiation of OPC into oligodendrocytes. We also found that ginkgolide B ameliorates oligodendrocytes apoptosis. Furthermore, ginkgolide B enhanced the expression of phosphorylated Akt and CREB. In conclusion, our data firstly show that ginkgolide B promotes oligodendrocyte genesis and oligodendrocyte myelin following a WML, possibly involving the Akt and CREB pathways.

Whole-body vibration (WBV), which is widely used as a type of exercise, involves the use of vibratory stimuli and it is used for rehabilitation and sports performance programmes. This study aimed to investigate the effect of WBV treatment in a chronic pain model after 10 WBV sessions. An animal model (chronic pain) was applied in 60 male Wistar rats (±180 g, 12 weeks old) and the animals were treated with low intensity exercise (treadmill), WBV (vibrating platform), and a combined treatment involving both. The controls on the platform were set to a frequency of 42 Hz with 2 mm peak-to-peak displacement, g ≈ 7, in a spiral mode. Before and after the vibration exposure, sensitivity was determined. Aβ-fibers-mediated mechanical sensitivity thresholds (touch-pressure) were measured using a pressure meter. C-fibers-mediated thermal perception thresholds (hot pain) were measured with a hot plate. After each session, WBV influenced the discharge of skin touch-pressure receptors, reducing mechanical sensitivity in the WBV groups (P < 0.05). Comparing the conditions “before vs. after”, thermal perception thresholds (hot pain) started to decrease significantly after the third WBV session (P < 0.05). WBV decreases mechanical hyperalgesia after all sessions and thermal sensitivity after the third session with the use of WBV.

Overdose of acetaminophen (APAP) is responsible for the most cases of acute liver failure worldwide. Hepatic mitochondrial damage mediated by neuronal nitric oxide synthase- (nNOS) induced liver protein tyrosine nitration plays a critical role in the pathophysiology of APAP hepatotoxicity. It has been reported that pre-treatment or co-treatment with glycyrrhizin can protect against hepatotoxicity through prevention of hepatocellular apoptosis. However, the majority of APAP-induced acute liver failure cases are people intentionally taking the drug to commit suicide. Any preventive treatment is of little value in practice. In addition, the hepatocellular damage induced by APAP is considered to be oncotic necrosis rather than apoptosis. In the present study, our aim is to investigate if glycyrrhizin can be used therapeutically and the underlying mechanisms of APAP hepatotoxicity protection. Hepatic damage was induced by 300 mg/kg APAP in balb/c mice, followed with administration of 40, 80, or 160 mg/kg glycyrrhizin 90 min later. Mice were euthanized and harvested at 6 h post-APAP. Compared with model controls, glycyrrhizin post-treatment attenuated hepatic mitochondrial and hepatocellular damages, as indicated by decreased serum glutamate dehydrogenase, alanine aminotransferase, and aspartate aminotransferase activities as well as ameliorated mitochondrial swollen, distortion, and hepatocellular necrosis. Notably, 80 mg/kg glycyrrhizin inhibited hepatic nNOS activity and its mRNA and protein expression levels by 16.9, 14.9, and 28.3%, respectively. These results were consistent with the decreased liver nitric oxide content and liver protein tyrosine nitration indicated by 3-nitrotyrosine staining. Moreover, glycyrrhizin did not affect the APAP metabolic activation, and the survival rate of ALF mice was increased by glycyrrhizin. The present study indicates that post-treatment with glycyrrhizin can dose-dependently attenuate hepatic mitochondrial damage and inhibit the up-regulation of hepatic nNOS induced by APAP. Glycyrrhizin shows promise as drug for the treatment of APAP hepatotoxicity.

Ras gene mutation or overexpression can lead to tumorigenesis in multiple kinds of cancer, including glioma. However, no drugs targeting Ras or its expression products have been approved for clinical application thus far. Adenoviral gene therapy is a promising method for the treatment of glioma. In this study, the human glioma cell line U251 was co-cultured with recombinant adenovirus KGHV500, and the anti-tumor effects of KGHV500 were determined by MTT, scratch test, Transwell invasion, and apoptosis assays. Then, KGHV500 was delivered via the intravenous injection of CIK cells into glioma xenografts. Tumor volume, ki67 proliferation index, apoptosis levels, and anti-p21Ras scFv expression were tested to evaluate targeting ability, anti-tumor efficacy, and safety. We found that the KGHV500 exhibited anti-tumor activity in U251 cells and increased the intracellular expression of anti-p21Ras scFv compared with that in the control groups. CIK cells delivered KGHV500 to U251 glioma cell xenografts and enhanced anti-tumor activity against glioma xenografts compared to that produced by the control treatment. In conclusion, targeting Ras is a useful therapeutic strategy for gliomas and other Ras-driven cancers, and the delivery of anti-p21Ras scFv by recombinant adenovirus and CIK cells may play an essential role in the therapy of Ras-driven cancers.