Placental expression of striatin & endothelial nitric oxide synthase in women with & without pre-eclampsia.

Background & objectives: Striatin is a multi-domain scaffolding protein essential for activating endothelial nitric oxide synthase (eNOS). However, its role in pre-eclampsia remains use explored. Hence, this study aimed to investigate the association between striatin and eNOS in regulating nitric oxide (NO) production in the placenta of women with and without pre-eclampsia. Methods: Forty pregnant women each without (controls) and with pre-eclampsia (cases) were enrolled in the study. Blood striatin and NO concentrations were detected by the ELISA. Protein expression of striatin, phosphorylated eNOS (peNOS), inducible NOS (iNOS) and phosphorylated NF-κB were measured in the placental tissues by Western blot. Twenty four hour urinary protein and serum urea, uric acid and creatinine were analyzed as an autoanalyzer. Placental histology was analyzed by haematoxylin and eosin staining. Results: Compared to normotensive pregnant women, the levels of serum NO and striatin were decreased in pre-eclamptic women. The protein expression of striatin and peNOS was significantly reduced (P<0.05) while p65NF-κB and iNOS were upregulated considerably (P<0.05) in the placenta of cases compared to controls. Interpretation & conclusions: Our results show for the first time that decreased striatin expression was associated with decreased peNOS protein expression in the placental tissue of pre-eclamptic women. Interestingly, no significant difference was found in blood striatin or NO levels between controls and cases. Thus, therapies that improve placental striatin expression are attractive possibilities, both for prevention as well as treatment of endothelial dysfunction in pre-eclampsia.

Role of zonula occludens in gastrointestinal and liver cancers.

A growing body of evidence suggests that tight junction (TJ) proteins play a crucial role in the pathogenesis of various diseases, including gastrointestinal (GI) cancer and hepatocellular carcinoma (HCC). TJ proteins primarily maintain the epithelial and endothelial cells intact together through integral proteins however, recent reports suggest that they also regulate gene expression necessary for cell proliferation, angiogenesis, and metastasis through adapter proteins such as zonula occludens (ZO). ZO proteins are membrane-associated cytosolic scaffolding proteins that modulate cell proliferation by interacting with several transcription factors. Reduced ZO proteins in GI cancer and HCC are correlated with tumor development and poor prognosis. Pubmed has searched for using the keyword ZO and gastric cancer, ZO and cancer, and ZO and HCC for the last ten years to date. This review summarized the role of ZO proteins in cell proliferation and their expression in GI cancer and HCC. Furthermore, therapeutic interventions targeting ZO in GI and liver cancers are reviewed.

Nimbolide attenuates gut dysbiosis and prevents bacterial translocation by improving intestinal barrier integrity and ameliorating inflammation in hepatocellular carcinoma.

Gut microbiota imbalance plays a key pathological role in hepatocellular carcinoma (HCC) progression; however, the mechanism is poorly understood. We previously showed nimbolide impede tumor development by improving hepatic tight junction (TJ) proteins expression and attenuating inflammation in HCC mice. Here, we aimed to study the role of nimbolide in regulating gut microbiota imbalance and bacterial translocation (BT) through modulating intestinal TJ proteins in an experimental hepatocarcinogenesis. Nimbolide (6 mg/kg) was administered orally for 4 weeks following induction of HCC in mice at the 28th week. Nimbolide treatment attenuated the gut microbiota imbalance by decreasing 16 s rRNA levels of Escherichia coli, Enterococcus, Bacteroides and increasing Bifidobacterium, and Lactobacillus in the intestinal tissue, which was otherwise altered in HCC mice. Furthermore, nimbolide improved intestinal barrier integrity in HCC mice by upregulating TJ proteins such as occludin and ZO-1 expression and subsequently prevented hepatic BT and decreased BT markers such as LBP, sCD14, and procalcitonin in the plasma of HCC mice. Moreover, nimbolide ameliorated intestinal and hepatic inflammation by downregulating TLR4, MyD88, and NF-κB protein expression in HCC mice. Thus, nimbolide represents a novel therapeutic drug for HCC treatment by targeting the gut-liver axis, which plays an imperative role in HCC pathogenesis.

Nimbolide inhibits tumor growth by restoring hepatic tight junction protein expression and reduced inflammation in an experimental hepatocarcinogenesis.

BACKGROUND: Altered tight junction (TJ) proteins are correlated with carcinogenesis and tumor development. Nimbolide is a tetranotriterpenoid that has been shown to have antioxidant and anti-proliferative properties; however, its anticancer effects and molecular mechanism in hepatocellular carcinoma (HCC) remains obscure. AIM: To investigate the effect of nimbolide on TJ proteins, cell cycle progression, and hepatic inflammation in a mouse model of HCC. METHODS: HCC was induced in male Swiss albino mice (CD-1 strain) by a single intraperitoneal injection of 100 mg/kg diethylnitrosamine (DEN) followed by 80 ppm N-nitrosomorpholine (NMOR) in drinking water for 28 wk. After 28 wk, nimbolide (6 mg/kg) was given orally for four consecutive weeks in DEN/NMOR induced HCC mice. At the end of the 32nd week, all the mice were sacrificed and blood and liver samples were collected for various analyses. Macroscopic examinations of hepatic nodules were assessed. Liver histology and HCC tumor markers such as alpha-fetoprotein (AFP) and glypican-3 were measured. Expression of TJ proteins, cell proliferation, and cell cycle markers, inflammatory markers, and oxidative stress markers were analyzed. In silico analysis was performed to confirm the binding and modulatory effect of nimbolide on zonula occludens 1 (ZO-1), nuclear factor of kappa light polypeptide gene enhancer in B-cells (NF-κB), and tumor necrosis factor alpha (TNF-α). RESULTS: We found nimbolide treatment at a concentration of 6 mg/kg to HCC mice reduced hepatic tumor size by 52.08% and tumor volume (P < 0.01), and delayed tumor growth in HCC mice with a concomitant reduction in tumor markers such as AFP levels (P < 0.01) and glypican-3 expression (P < 0.05). Furthermore, nimbolide treatment increased tight junction proteins such as ZO-1 and occludin expression (P < 0.05, respectively) and reduced ZO-1 associated nucleic acid binding protein expression (P < 0.001) in HCC mice liver. Nimbolide treatment to HCC mice also inhibited cell proliferation and suppressed cell cycle progression by attenuating proliferating cell nuclear antigen (P < 0.01), cyclin dependent kinase (P < 0.05), and CyclinD1 (P < 0.05) expression. In addition, nimbolide treatment to HCC mice ameliorated hepatic inflammation by reducing NF-κB, interleukin 1 beta and TNF-α expression (P < 0.05, respectively) and abrogated oxidative stress by attenuating 4-hydroxynonenal expression (P < 0.01). Molecular docking studies further confirmed that nimbolide interacts with ZO-1, NF-κB, and TNF-α. CONCLUSION: Our current study showed for the first time that nimbolide exhibits anticancer effect by reducing tumor size, tumor burden and by suppressing cell cycle progression in HCC mice. Furthermore, nimbolide treatment to HCC mice ameliorated inflammation and oxidative stress, and improved TJ proteins expression. Consequently, nimbolide could be potentially used as a natural therapeutic agent for HCC treatment, however further human studies are warranted.

Increased systemic zonula occludens 1 associated with inflammation and independent biomarker in patients with hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is a frequent type of primary liver cancer, and its prevalence is increasing worldwide. Indeed, the underlying molecular mechanism is not well understood. Previous studies have shown evidence that tight junction (TJ) components were correlated with carcinogenesis and tumor development. Our aims were to determine the serum levels of tight junction protein Zonula Occludens (ZO)-1 and an inflammatory marker such as high-sensitive C-reactive protein (hs-CRP) in HCC patients compared to healthy volunteers and also to identify the association between ZO-1 and inflammation in HCC. METHODS: Thirty HCC patients and 30 healthy volunteers were recruited in the current study. Clinical data regarding child class, BCLC staging, the number of lesions, tumor size, absence or presence of metastasis, cirrhosis and hepatitis infection were also collected in HCC patients. Plasma ZO-1 and serum hsCRP were analyzed by EIA and ELISA respectively and biochemical parameters by autoanalyser (AU680 Beckman Coulter, USA). Furthermore, hepatic ZO-1 protein expression and tissue localization were examined. RESULTS: Compared to healthy individuals, the serum levels of bilirubin, alanine aminotransferase (ALT), aspartate aminotransferase (AST), gamma-glutamyltransferase (GGT) and alkaline phosphatase (ALP) were elevated significantly (P < 0.0001) whilst serum albumin level was significantly (P < 0.0001) decreased in HCC patients. Furthermore, tight junction protein ZO-1 concentration was significantly elevated in HCC patients compared to control subjects (648 ± 183.8 vs. 396.4 ± 135.8 pg/ml, respectively; P < 0.0001). Serum hsCRP level was also significantly increased in HCC patients compared to control subjects (17.25 ± 3.57 vs. 5.54 ± 2.62 mg/L, respectively; P < 0.0001). Moreover, decreased protein expression of ZO-1 was found in liver tissue obtained from HCC patients. CONCLUSION: Our findings show for the first time that the systemic concentration of ZO-1 was significantly elevated in HCC patients and is positively correlated with inflammatory markers. Thus, the current study showing evidence that inflammation promotes plasma ZO-1 concentration and raises the possibility that it could be used as a potential diagnostic biomarker for HCC progression.