BACKGROUND:Calcium citrate has a better solubility than calcium phosphate, calcium sulfate, and other calcium biomaterials. The synthetic calcium citrate has a good denseness, and stably releases calcium ions at a high efficiency during the degradation. Consequently, it may be more suitable for the fil ing of fracture defects, providing needed calcium ions for early fracture healing.
OBJECTIVE:To prepare calcium citrate biomaterials with a novel formulation based on the natural bio-mineralized oyster shel s and citric acid so as to expect to get a good application in fracture healing repair.
METHODS:Crushing, grinding, and chemical reaction methods were used for refinement. Particle size analyzer, X-ray diffraction, scanning electron microscope, and Fourier transform infrared spectroscopy were adopted for analysis of the size distribution, composition, mineral phases, and micro-morphology. Biological characteristics were evaluated through a simulated body fluid experiment.
RESULTS AND CONCLUSION:Oyster shel powder was reacted with saturated citric acid to produce the calcium citrate material that had uniform crystal structure and compact bonding among crystal bodies, and exhibited a certain mechanical ability. The calcium citrate material had a good crystal structure that was conductive to prolong the degradation time. The calcium citrate released calcium ions slowly, and did not produce dramatic changes in the pH value (7.20-7.46) of the surrounding in the dissolution process. With the gradual degradation of calcium citrate materials, Ca2+concentration in solution increased gradual y and stably, and ultimately achieved an appropriate concentration of 7 mmol/L, suitable for osteoblast proliferation and differentiation. Calcium citrate prepared using natural oyster shel has good biological properties, and exhibits a natural superiority to artificial bone materials.

Objective To prepare a detective membrane strip for detection of food allergen-specific IgE in serum samples and estimate its clinical application value in allergic diseases. Methods The crude extracts of the food allergens were prepared. Nitrocellulose membrane as the solid support was selected and the coating and the detecting conditions were optimized. The membrane strips were used to detect serum samples in 210 patients with allergic diseases and the results were compared with German Allergy Screentesting system. Results The optima] experimental conditions were as follows: The NC membrane was adopted as the solid support. After being spotted, the food allergens were incubated for 2 hours at room temperature, followed by 2% PVA blocking for 1 hour. After serum samples were diluted (1: 10) and incubated for 2 hours at room temperature, the concentration of anti-human IgE was 2 μg/mL Compared with the German Allergy Screen-testing system, their positive detectical coincidence was 63.6%, and negative detectical coincidence was 94. 6%. The two methods had no difference in detecting the majority of food allergens such as egg white, milk, peanut, soybean, crab and shrimp (X2 2.53, 2.40, 2.08, 2.38, 0.17,1.13, P>0.05). Conclusions The advantages of our method for detecting allergic diseases are little serum needed, multiple detective allergens, simple manipulation and low cost. This method has obvious clinical application value, which should be a new detective method for the allergic diseases with broad perspectives.

Objective To observe the effect of JLP on transdifferentiation of human renal proximal tubular epithelial cells (HK-2),and to investigate the role of p38 MAPK signaling pathway in this process.Methods The knock-down plasmids of JLP were constructed.HK-2 cells were randomly divided into four groups:negative control cells (Ctrl-shRNA group),knock-down jlp cells (jlpshRNA group),negative control cells with FGF-2 treatment (FGF-2 group) and knock-down jlp cells with FGF-2 treatment(jlp-shRNA +FGF-2 group).The expressions of JLP,E-cadherin,TGF-β1,α-SMA,p-p38 MAPK protein were detected by Western blotting.After the induction of FGF-2 for 24 hours,the expressions of α-SMA,COL-Ⅰ,FN were detected by immunocytochemistry.Results Compared with Ctrl-shRNA group,the expression of JLP protein was significantly down-regulated in FGF-2 group.Compared with FGF-2 group,the expressions of TGF-β1,α-SMA,p-p38 MAPK protein were significantly up-regulated,while E-cadherin protein was significantly down-regulated (P ＜ 0.05).Compared with FGF-2 group,the expressions of α-SMA,COL-Ⅰ,FN immunostaining increased markedly in jlp-shRNA+FGF-2 group.Conclusion Scaffolding protein JLP is critical in preventing EMT in the course of fibrosis through the inhibition of p-p38 activation in HK-2 cells.

Objective To observe the effect of JLP deficiency on the progression of renal interstitial fibrosis in mice model of unilateral ureteral obstruction (UUO),and to investigate the role and underlying mechanism of JLP in the development of renal fibrosis in obstructive nephropathy.Methods jlp Wild type (jlp+/+) and jlp deficient (jlp-/-) mice were divided into four groups:jlp+/+-and jlp-/--sham-operated groups(jlp-/--sham group and jlp+/+-sham group),jlp+/+-and jlp-/--unilateral ureteral obstruction (UUO)-operated groups (jlp/--UUO group and jlp+/+-UUO group).Mice were sacrificed at 7 days and 14 days after the operation respectively to evaluate the fibrosis by Masson staining.The expression of JLP in jlp +/+ renal tissue was assayed by immunohistochemistry staining,immunofluorescence and Western blotting.Immunohistochemical staining was used to detect the expression of α-smooth muscle actin (α-SMA),collagen Ⅰ (COL-Ⅰ),collagen Ⅲ (COL-Ⅲ) and transforming growth factor-β1 (TGF-β1) in sham and UUO groups.Besides,the α-SMA,COL-Ⅰ,COL-Ⅲ,TGF-β1,p-Smad2 and p-Smad3 protein levels were also analyzed by Western blotting in four groups.Results The expression of JLP was mainly demonstrated in the renal tubules of mice.A large amount of collagen deposition was observed in the renal interstitial area in jlp-/--UUO group compared to jlp+/+-UUO group.Similarly,the expression of α-SMA,COL-Ⅰ,COL-Ⅲ and TGF-β1 was significantly increased in the kidney cortices in jlp-/--UUO-operated groups.Meanwhile,Western blotting showed that the expression of α-SMA,COL-Ⅰ,COL-Ⅲ,and TGF-β1 protein was obviously higher in jlp-/--UUO group.Moreover,the expression of p-Smad2 and p-Smad3 protein was markedly higher in jlp-/--UUO group.Conclusion Scaffolding protein JLP is critical in preventing renal fibrosis through the inhibition of TGF-β1 expression and myo-fibroblast production.

Objective: To investigate the effect of enolase 1 (ENO1) expression on proliferation, apoptosis and migration of lung cancer PC14 cells. Methods: ENO1 over-expression vector-pcDNA3.1/ENO1 was constructed and transfected into PC14 cells at logarithmic growth phase with liposome LipofectamineTM 2000. G418 was used to screen PC14 cells that stably expressing ENO1. The effects of ENO1 over-expression on proliferation, migration and apoptosis of PC14 cells were detected by CCK-8 method, scratch-healing assay and flow cytometry, respectively. Results: The ENO1 over-expression cell model was successfully constructed. Compared with PC14-vehicle and wild-type PC14 cells, the mRNA and protein expression levels of ENO1 in PC14-ENO1 cells were significantly elevated (all P<0.05), and the proliferation of PC14-ENO1 cells was significantly increased (all P<0.05). The relative mobility of PC14ENO1 cells was significantly higher than that of pcDNA3.1-vehicle cells and wild-type PC14 cells ([13.26±1.13]% vs [8.46±1.11]%, [7.86±1.00]%, both P<0.05). There was no significant difference in apoptotic rate among PC14-ENO1, PC14-vehicle and PC14 cells (all P> 0.05) Conclusion: Over-expression of ENO1 promotes proliferation and migration of lung cancer PC14 cells.

Objective To investigate the roles of A kinase anchoring protein1(AKAP1)in high-glucose induced mitochondrial fission in podocytes.Methods Conditionally immortalized human podocytes were cultured in serum-free medium for 24 hours,and then exposed to different glucose concentration conditions in different time periods.The protein expressions of AKAP1 were observed by immunofluorescence,and AKAP1,dynamin related protein1 (Drp1) and phospho Ser 637-Drp1 (p-Drp1)were analyzed by Western blotting.AKAP1 siRNA was transfected to block AKAP1 expression.Podocytes were then divided into normal control group (5 mmol/L glucose),hypertonic group (30 mmol/L mannitol+5 mmol/L glucose),high glucose group (35 mmol/L glucose),and high glucose+AKAP1 siRNA group.Mitochondrial morphological changes were assessed by mitotracker red staining.Podocyte apoptosis was assessed by flow cytometry.Results Compared with normal group,high-glucose induced more podocytes apoptosis (P ＜ 0.05),more mitochondrial fission with decreased aspect ratio and form factor (all P ＜ 0.05).Upregulated AKAP1 protein level,and increased ratio of p-Drp1/Drp1 (all P ＜ 0.05) in time and concentration dependent manners were also obscrvcd.Compared with high glucose group,transfection of AKAP1 siRNA showed less apoptosis (P ＜ 0.05),less mitochondrial fission with increased aspect ratio and form factor (all P ＜ 0.05),and down-regulated AKAP1 protein level as well as p-Drp1/Drp1 ratio (all P ＜ 0.05).Conclusion High glucose induced mitochondrial fission might be induced through AKAP1-Drp1 pathway.

Objective:To investigate the role of mitofusion 2 (Mfn2) in high glucose (HG)-induced endoplasmic reticulum stress (ERS) and apoptosis of podocytes.Methods:(1) Streptozocin was used to induce a diabetes mellitus (DM) rat model. Renal histopathological changes in rats were observed by HE staining. Expression of Mfn2 and CCAAT/enhancer-binding protein homologous protein (CHOP) in glomeruli was observed by immunohistochemistry. Protein levels of Mfn2, protein kinase RNA-like ER kinase (PERK), phospho(p)-PERK, and CHOP in glomeruli were analyzed by Western blotting. (2) Conditionally immortalized human podocytes (HPC) cultured in vitro were divided into control, mannitol (MA) and HG groups. Expression of Mfn2 was observed by immunofluorescence. Protein levels of Mfn2, p-PERK, PERK and CHOP in HPC were analyzed by Western blotting. Podocyte apoptosis in each group was evaluated by flow cytometry with AnnexinⅤ-PE/7AAD double staining method. (3) HPC were divided into control, HG, HG+Mfn2-Myc plasmid-transfected and HG+control plasmid-transfected groups. Protein levels of Mfn2, p-PERK, PERK and CHOP in HPC were analyzed by Western blotting. Expression of CHOP was observed by immunofluorescence. Mitochondrial membrane potential in each group was observed by mitochondrial membrane potential assay kit with JC-1. Podocyte apoptosis in each group was evaluated by flow cytometry with AnnexinⅤ-PE/7AAD double staining method. Results:(1) Compared with the control group, the glomerular mesangial matrix of the DM group rats was significantly proliferated, and the expression of Mfn2 was down-regulated with the expression of ERS-related proteins p-PERK/PERK and CHOP up-regulated (all P<0.05). (2) Compared with the control group, Mfn2 was down-regulated and p-PERK/PERK and CHOP were up-regulated in HPC of HG group (all P<0.05). Apoptosis of HPC was also increased in HG group. There was no significant difference in the above indicators between the control group and the mannitol group (all P>0.05). (3) Compared with the HG group, mitochondrial membrane potential of HPC was alleviated and apoptosis of HPC was decreased in HG+Mfn2-Myc plasmid-transfected group ( P<0.05). P-PERK/PERK and CHOP were down-regulated in HG+Mfn2-Myc plasmid-transfected group (both P<0.05). There was no significant difference in the above indicators between the HG group and the HG+control plasmid-transfected group (all P>0.05). Conclusions:Mfn2 down-regulation in HG-stimulated podocytes may induce ERS to increase apoptosis of podocytes. Up-regulation of Mfn2 can alleviate the HG-induced ERS and apoptosis in podocytes.

Objective: To investigate and analyze the drinking water quality among rural primary and secondary schools in Guanzhong area in 2018, and to provide a basis for the targeted improvement of water supply facilities. Methods: Develop a questionnaire on the basic situation of centralized water supply in rural schools, according to the Standard Test Method for Drinking Water (GB/T 5750—2006) and the Sanitary Standard for Drinking Water (GB 5749—2006) for the rural school network in Guanzhong area. The peripheral water collection and testing carried out single factor and multifactor statistical analysis on the relationship between water quality influencing factors and water quality pass rate. Results: The total qualified rate of drinking water quality in rural schools in Guanzhong area was 59.1%. Univariate analysis showed that water quality rate was affected by four factors including water source type, engineering type, sanitation permit and disinfection equipment use, and the difference was statistically significant(P<0.05). Unconditional two-class logistic regression analysis showed that disinfection (OR=3.14), engineering type (OR=2.05), and sanitation permit (OR=1.99)(P<0.05) affected the water quality pass rate. Conclusion It is recommended to further strengthen the investment in the renovation of water supply for rural schools in Guanzhong area, and specifically strengthen water supply treatment and standard disinfection.

Objective: To understand the status and trend of heavy metal indicators of drinking water in rural schools in different regions of Shaanxi Province, so as to provide scientific basis for safety of drinking water in rural schools. Methods: In 2017-2019, 697 rural school water supply projects in Shaanxi Province were tested for heavy metal indicators in the peripheral water. According to the sanitary standard for drinking water (GB 5749—2006), five heavy metal indicators, including arsenic, cadmium, hexavalent chromium, lead and mercury, were analyzed and evaluated in different years and regions. Results: A total of 2 298 valid water samples were collected and analyzed in 3 years. Except that lead and mercury are all up to standard, the standard rates of other heavy metals such as arsenic, cadmium and hexavalent chromium were 98.83%, 99.91% and 96.95% respectively. Compared with the Northern Shaanxi plateau and Qinba mountain area, the standard rate of water arsenic in Guanzhong Plain was lower (χ2=5.67, 13.59,P<0.01). The standard rate of hexavalent chromium was the highest in Qinba mountain area, followed by Guanzhong Plain, and the lowest in Northern Shaanxi plateau (χ2=20.48, 17.05, 48.32, P<0.01). Two samples of cadmium exceeding standard were from the Northern Shaanxi plateau. Conclusion The heavy metal index of drinking water in rural schools in Shaanxi Province exceeds the standard, which has obvious regional characteristics. We should focus on the harm of arsenic, hexavalent chromium and other heavy metals to the health of students in the Northern Shaanxi plateau and Guanzhong Plain. Cadmium and mercury in drinking water in local areas should be paid continued attention. Safety of drinking water in schools should be ensured from the aspects of water source selection and water treatment technology.

Liver cancer is one of the most common and fatal malignancies worldwide, with high mortality and morbidity. As an important tumor suppressor gene, p53 gene plays a key role in cell cycle regulation, cell growth and apoptosis of tumor cells. P53 gene mutations are closely related to the occurrence of liver cancer. At present, the treatment of liver cancer includes surgery, local treatment and systemic treatment. However, since most liver cancer patients are in advanced stages at the time of diagnosis, non-surgical treatment has become the best choice for the treatment of patients with intermediate and advanced liver cancer. But patients still have problems such as short survival and easy drug resistance. Therefore, the development of new treatment options is essential to improve the treatment outcomes of liver cancer patients. P53-related gene therapy products have great potential as drugs for the treatment of liver cancer. This article will briefly review the relationship between p53 gene and primary liver cancer, and explore its role in diagnosis and treatment, in order to provide a new direction for future clinical treatment.