Overexpression of miR-195-5p reduces osteoporosis through activating BMP-2/SMAD/Akt/RUNX2 pathway via targeting SMURF1.

Osteoporosis (OP) is among the most common frequent chronic metabolic bone diseases in postmenopausal women. Here, the effect and underlying mechanisms of miR-195-5p in OP were investigated both in vivo and in vitro. In this study, the microgravity (MG) environment was simulated in MC3T3-E1 cells, and miR-195-5p overexpression or SMURF1 knockdown model was constructed to test their effects on the proliferation, apoptosis and osteogenic differentiation of MC3T3-E1 cells. Furthermore, an OVX mouse model was constructed in vivo, and adenovirus-loaded miR-195-5p mimics were administered to the mice to overexpress miR-195-5p. HE staining and µCT were adopted to observe pathological changes of femur. The targeted relationship between miR-195-5p and SMURF1 was predicted by bioinformatics analysis and verified by the dual-luciferase reporter assay and RNA immunoprecipitation (RIP) experiment. The results indicated that miR-195-5p was down-regulated in the head of femur of OP mouse model and MC3T3-E1 cells subjected to MG microenvironment. In addition, overexpression of miR-195-5p promoted MC3T3-E1 cell osteogenic differentiation and inhibited apoptosis. Mechanistically, SMURF1 is identified as a target of miR-195-5p, and overexpressing miR-195-5p activates the BMP-2/SMAD/Akt/RUNX2 signal by inhibiting the SMURF1 expression. Moreover, SMURF1 downregulation accelerated the osteogenic differentiation of MC3T3-E1 cells and attenuated MG-mediated apoptosis. In addition, upregulating miR-195-5p reduced osteoporosis in the OVX mouse model, accompanied with SMURF1 downregulation and BMP-2/SMAD/Akt/RUNX2 pathway activation. Collectively, miR-195-5p enhances osteogenic differentiation of osteoclast and relieve OP progression in the mouse model through activation of the BMP-2/SMAD/Akt/RUNX2 axis by targeting SMURF1.

Optimization of venous return tubing diameter for cardiopulmonary bypass.

OBJECTIVE: To determine the optimal venous tubing diameter for adult cardiopulmonary bypass (CPB) to improve gravity drainage and to reduce priming volume. METHODS: (A) Maximum bovine blood flow rates by gravity drainage were assessed in vitro for four different tubing diameters (1/2, 3/8, 5/16,1/4 inch) with three different lengths and various pre- and afterloads. Based on the results of (A) and multiple regression analyses, we developed equations to predict tubing sizes as a function of target flows. (C) The equations obtained in (B) were validated by ex vivo bovine experiments. (D) The clinically required maximal flows were determined retrospectively by reviewing 119 perfusion records at Zurich University. (E) Based on our model (B), the clinical patient and hardware requirements, the optimal venous tubing diameter was calculated. (F) The optimized venous tubing was evaluated in a prospective clinical trial involving 312 patients in Hangzhou. RESULTS: For a mean body surface area of 1.83+/-0.2 m(2), the maximal perfusion flow rate (D) achieved with 1/2-inch (=1.27 cm(2)) venous tubing was 4.62+/-0.57 l/min (range: 2.50-6.24 l/min). Our validated model (B,C) predicted 1.0 cm(2) as optimal cross-sectional area for the venous line. New tubing packs developed accordingly were used routinely thereafter. The maximal flow rate was 4.93+/-0.58 l/min (range: 3.9-7.0) in patients with a mean body surface area of 1.62+/-0.21 m(2). CONCLUSION: The new venous tubing with 1.0-cm(2) cross-sectional area improves the drainage in the vast majority of adult patients undergoing CPB and reduces the priming volume (-27 ml/m). Reduced hemodilution can prevent homologous transfusions if a predefined transfusion trigger level is not reached.

Expression of insulin-like growth factor II and its receptor in liver cells from patients with chronic liver diseases.

AIM: To study the relationship between insulin-like growth factor II (IGF-II), IGF-II receptor, and chronic liver diseases and to investigate the clinical mechanisms of human hepatocellular carcinoma (HCC) development. METHODS: We analyzed IGF-II and IGF-II receptor poly (A)+ mRNA in dysplasia liver cell (DLC; n = 10), liver cirrhosis (LC; n = 9), and chronic active hepatitis (CAH; n = 9) specimens by Northern blot using human IGF-II and IGF-II receptor DNA probes labeled with (32) P through nick translation. RESULTS: Expression of IGF-II in DLC samples (10/10, 100%) was higher than in CAH (3/9, 33%) and LC samples (3/9, 33%) (P < 0.01). Expression of IGF-II receptor in DLC samples (7/10, 70%) was significantly higher than in CAH (2/9, 22%) and LC samples (3/9, 33%). Data on hepatitis B virus (HBV) infection status from different chronic liver disease samples were also analyzed. CONCLUSION: Overexpression of IGF-II and IGF-II receptor in DLC samples was associated with a preceding step to malignant phenotype hepatocyte transformation and may be of diagnostic value for early detection of hepatocellular carcinoma (HCC). Persistent HBV infection was strongly associated with abnormal IGF-II and IGF-II receptor mRNA expression, suggesting that an autocrine or paracrine mechanism is involved in the regulation of growth in liver cell carcinogenesis.

Studies on the hypomethylation of c-myc, c-Ha-ras oncogenes and histopathological changes in human gastric carcinoma.

In order to study the status of DNA methylation of specific oncogenes and the relationship between them and the pathological changes in gastric carcinoma, we analysed the methylated status of c-myc, c-Ha-ras oncogenes by Southern blot hybridization. Genomic DNA from cancerous, paracancerous and non-cancerous areas of surgically resected specimens were examined in 22 cases of advanced human gastric carcinoma. Specimens were digested by the restriction endonucleases MspI/HpaII, which are able to cleave between methylated and non-methylated cytosine at their nucleotide recognition site the DNA 5'-CCGG sequence, and were hybridized with c-myc, c-Ha-ras oncogene probes. Moreover, the corresponding pathological changes in gastric carcinoma were observed. The results showed that c-myc, c-Ha-ras oncogenes from cancerous (10/22, 5/10) and paracancerous areas (13/22, 4/10) were hypomethylated and that there was no significant relationship between them and the histopathological changes.

Transthyretin (prealbumin) gene in human primary hepatic cancer.

From a subtracting cDNA library constructed from normal liver versus human primary hepatic cancer (PHC) a cDNA clone pG8 was isolated. Using it as a probe, RNA extracted from one human liver and 9 PHC samples were analyzed by Northern hybridization. As expected, its mRNA was highly expressed in liver; however, the expression was strikingly suppressed in PHC. Only weak signal was observed in 2 out of 9 PHC, while no signal was detectable in the other 7 samples. Utilizing pG8 as a probe, DNA from the same PHC specimens was analyzed after MspI digestion and Southern hybridization. Deletion of DNA fragment was observed in 4 out of 9 samples. In further study of cancer and non-cancerous liver from other 7 PHC patients, similar deletion of DNA fragments in cancer was observed in 4 out of 7 samples. After sequencing of the clone of 572 bp, it was unexpectedly found that pG8 was completely homologous to the coding sequence of transthyretin, TTR gene, as TTR (or prealbumin) gene has been known to be linked to a hereditary disorder, familial amyloidosis (FAP), and related to thyroxine transport and binding to retinol-RBP (the retinol binding protein) complex. This is the first report of a study on TTR in human primary hepatic cancer. Since TTR gene was strikingly suppressed in mRNA expression and possibly defective in its gene structure, it was strongly implicated that TTR might be an important gene marker or a candidate of anti-oncogene for human PHC. The biological activity of TTR gene is under study.

Chromium-induced kidney disease.

Kidney disease is often cited as one of the adverse effects of chromium, yet chronic renal disease due to occupational or environmental exposure to chromium has not yet been reported. Occasional cases of acute tubular necrosis (ATN) following massive absorption of chromate have been described. Chromate-induced ATN has been extensively studied in experimental animals following parenteral administration of large doses of potassium chromate (hexavalent) (15 mg/kg body weight). The chromate is selectively accumulated in the convoluted proximal tubule where necrosis occurs. An adverse long-term effect of low-dose chromium exposure on the kidneys is suggested by reports of low molecular weight (LMW) proteinuria in chromium workers. Excessive urinary excretion of beta 2-microglobulin, a specific proximal tubule brush border protein, and retinol-binding protein has been reported among chrome platers and welders. However, LMW proteinuria occurs after a variety of physiologic stresses, is usually reversible, and cannot by itself be considered evidence of chronic renal disease. Chromate-induced ATN and LMW proteinuria in chromium workers, nevertheless, raise the possibility that low-level, long-term exposure may produce persistent renal injury. The absence of evidence of chromate-induced exposure may produce persistent renal injury. The absence of evidence of chromate-induced chronic renal disease cannot be interpreted as evidence of the absence of such injury. Rather, it must be recognized that no prospective cohort or case-control study of the delayed renal effects of low-level, long-term exposure to chromium has been published.