Study on the mechanism of miR-23b-3p regulating osteogenic differentiation of renal interstitial fibroblasts and participating in Randall's plaque formation / 安徽医科大学学报

Objective To explore the effect of miR-23b-3p regulation on osteogenic differentiation of renal intersti-tial fibroblasts(hRIFs)on the formation of Randall plaque and its possible mechanism.Methods qRT-PCR was used to detect the expression levels of miR-23b-3p and osteogenic marker:myocyte enhancer factor 2C(MEF2C),osteocalcin(OCN),osteopontin(OPN),runt-related transcription factor 2(Runx2)mRNA in Randall plaque tis-sue of CaOx stone patients(RP)and normal papillary tissue of kidney tumor patients undergoing nephrectomy(nRP).Isolation and culture of human normal hRIFs were isolated and cultured in vitro.The miR-23b-3p overex-pression plasmid pSi-miR-23b-3p and its negative no-load plasmid pSi-NC,the MEF2C lentivirus overexpression plasmid Lv-MEF2C and the no-load plasmid Lv-NC were transfected into hRIFs cells,and the cells were induced to osteogenic differentiation for 14 days.The activity of alkaline phosphatase(ALP)was determined by ELISA.Aliz-arin red staining was used to observe the formation of mineralized nodules.The expression levels of miR-23b-3p and MEF2C,OCN,OPN,Runx2 mRNA were detected by qRT-PCR.The expression level of MEF2C protein was de-tected by Western blot.Dual luciferase reporter gene assay verified the targeting relationship between miR-23b-3p and MEF2C.Results ① Compared with the nRP group,miR-23b-3p was low expressed and MEF2C,OCN,OPN,and Runx2 were highly expressed in the RP group.② 14 days after osteogenic induction of hRIFs cells,the activity of ALP in cells significantly increased,the ability of cells to form mineralized nodules was enhanced,the expression level of miR-23b-3p significantly decreased,the mRNA expression levels of MEF2C,OCN,OPN,and Runx2 significantly increased,and the expression level of MEF2C protein significantly increased.③ Overexpres-sion of miR-23b-3p decreased the activity of ALP in hRIFs cells after osteogenic induction,inhibited the formation of mineralized nodules in cells,and down-regulated the mRNA expression levels of OCN,OPN,and Runx2 in cells.④ Overexpression of MEF2C reversed the inhibitory effect of miR-23b-3p overexpression on osteoblast differ-entiation of hRIFs cells.⑤ MEF2C was the downstream target gene of miR-23b-3p.Conclusion miR-23b-3p is underexpressed in RP tissues and during osteoblastic differentiation of hRIFs cells.Up-regulation of miR-23b-3p in-hibits osteogenic differentiation of hRIFs cells,and its mechanism may be related to targeted silencing MEF2C.

Comparative Analysis of the Hospitalization Expenses Before and After the City-level Public Hospital Reform: Taking a Third-level Grade-A Hospital in Anhui Province as an Example / 中国医院管理

Objective To understand the present situation of city-level public hospital reform and to analyze the impacts on different clinical departments by analyzing the changes of hospitalization expenses of a third-level grade-A hospital from Anhui Province.Methods The inpatient data from April to September,2014 and 2015 of the third-level grade-A hospital were collected.Three typical diseases in Department of Internal Medicine,Surgery,Gynecology and Pediatrics respectively,were chosen to compare the changes of expenses and the structure before and after the reform.Results Overall,hospitalization expenses saw a declining trend after city-level public hospital reform.But,it was different in each department.The structure of hospitalization expenses became more reasonable after reform,however,the medical material expense remained a question.Conclusion The city-level public hospital reform reduces drug costs and inspection fees for inpatients effectively,thereby abating the overall cost of hospitalization,and the structure of hospitalization expenses becomes more reasonable.But the changes of hospitalization expenses in different departments reflect some problems.

The effect of obstructive sleep apnea syndrome on vascular endothelial function and cardiovascular events in elderly patients / 中华老年医学杂志

Objective To retrospectively analyze the cardiovascular outcomes of the elderly patients with obstructive sleep apnea syndrome (OSAS) and the influence factors. Methods According to polysomnography examination, 79 OSAS patients and 60 patients without OSAS were selected and divided into 3 groups: elder OSAS group [39 patients older than 65 years, respiratory apnea index (AHI)≥5], non-elder OSAS group (40 patients less than 65 years old, AHI≥5) and elder control group (60 patients older than 65 years, AHK5). All patients were followed up by telephone and clinic consulting. The median follow-up duration was 25 months. All patients received the following studies: (1)Flow-mediated vasodilation (FMD), which was assessed by high-resolution ultrasound technique. (2)The sleep-related breathing events and serum biochemical indexes. (3)The death due to cardiovascular disease (CVD), angina, myocardial infarction and stroke. Results (1) FMD was significantly lower in elder OSAS group than in elder control group (P<0. 01). (2)In elder OSAS group versus elder control group, BMI was significantly higher (P<0. 01), while both lowest pulse oxygen saturation (LSpO_2 ) and mean series pulse oximeter ( MeanSpO_2 ) were significantly lower (P< 0.01 and P<0. 05, respectively). Multiple logistic regression analysis showed that impaired fasting plasma glucose was the primary injury factor for FMD (OR=1. 83, 95% CI:1. 11～3.03), and LSpO_2 was the secondary injury factor (OR = 0.92, 95% CI: 0. 85～1. 00). (3) The incidence of cardiovascular events in the 3 groups: the incidence of cardiovascular events was significantly higher in elder OSAS group than in the other 2 groups (χ~2= 7. 339, P<0. 05). Multiple logistic regression analysis showed that FMD (OR=1. 33, 95% CI:1. 06～1. 66)and hs-CRP (OR = 0.51, 95% CI: 0. 34～0.76) were closely related with prognosis. Conclusions Compared with non-elder OSAS group and elder control group, vascular endothelial function impairment is more serious and the incidence of cardiovascular events is higher in elder OSAS group. So OSAS may influence the prognosis of the patients by injuring the vascular endothelial function.

A novel method to evaluate myocardial injury: cloning of anti-cTnI murine antibody Fab fragment and DNA sequence analysis / 中国组织工程研究

BACKGROUND: To apply mouse anti-human cTnI monoclonal antibody as the drug vector in the treatment and diagnosis of myocardial injury, it is important to degrade the immunity of murine antibody and overcome human anti-mouse reaction. Humanization has been applied as an attempt to resolve this problem.OBJECTIVE: To clone murine anti-cTnI Fab fragment and analyse the nucleotide and deduced amino acid sequences.DESIGN: Single sample study.SETTING: An institute of cardiovascular disease under a medical university-affiliated hospitalMATERIALS: The study was conducted in the Institute of Cardiovascular Diseases, First Affiliated Hospital of Nanjing Medical University from January 2003 to May 2004. The hybridoma cell line JS200202 which secrets the anti-cTnI monoclonal antibody was provided by Institute of Cardiovascular Disease, First Affiliated Hospital of Nanjing Medical University.METHODS: IgG heavy chain primers and κ light chain primers of amplified mouse were designed. Total RNA was extracted from hybridoma cells which secrete cTnI. Reverse transcription polymerase chain reaction(RT-PCR) was amplified. Cloning and subsequent sequence analysis of the Fab fragment was performed. The deduced amino acid sequence was compared and analysed with previously published sequences.MAIN OUTCOME MEASURES: Heavy chain Fd segment and κ light chain gene sequence and its subgroups.RESULTS: A band of approximate 700 and 800 base pairs were amplified using IgG heavy chain primers and κ light chain primers respectively. Sequence analysis indicated that the deduced amino acid sequences were in consistent with the characterization of the amino acid in the murine IgGl Fab fragment(GenBank accession NO AY484430, AY484431; Protein Bank accession NO AAR83243, AAR83244).CONCLUSION: A complete murine anti-cTnI Fab fragment was obtained in this study, which may provide basis for the production of the chimeric anti-cTnI antibody.

Cloning and Identification of the Gene Fragments of Paragonimus westermani / 中国寄生虫学与寄生虫病杂志

Objective To screen and identify the recombinants from the cDNA library of the adult Paragonimus west-ermani (PwA) for immunodiagnosis and immunoprophylaxis. Methods PwA cDNA library was screened with the PwA antigen immunized rabbit sera(IRS) pre-absorbed by the extract of E. coli XLl-Blue. The recombinants from positive clones were amplified by PCR, sequenced and cut off by KpnI/BarnHI and, then sub-cloned into pRESETB vector. The fusion protein was expressed,analysed by SDS-PAGE and identified by Western blotting with immune rabbit serum against worm antigen of Paragonimus westerrnani. Results The inserted cDNA fragment from the positive clone Pw-2 was about 800 bp, which contained an open reading frame(ORF) encoding Pw pre-procathepsin L belonging to cysteinase family. Expression product of Pw-2 was a fusion protein of 32 kDa, which can be recognized by immune rabbit serum against worm antigen of Paragonimus 晈esterrnani. Conclusion A recombinant plasmid Pw-2 encodes Pw pre-procathepsin L is constructed.