Artificial cavernosa-like tissue based on multibubble Matrigel and a human corpus cavernous fibroblast scaffold.

Ex vivo tissue culture of the human corpus cavernosum (CC) can be used to explore the tissue structural changes and complex signaling networks. At present, artificial CC-like tissues based on acellular or three-dimensional (3D)-printed scaffolds are used to solve the scarcity of primary penis tissue samples. However, inconvenience and high costs limit the wide application of such methods. Here, we describe a simple, fast, and economical method of constructing artificial CC-like tissue. Human CC fibroblasts (FBs), endothelial cells (ECs), and smooth muscle cells (SMCs) were expanded in vitro and mixed with Matrigel in specific proportions. A large number of bubbles were formed in the mixture by vortexing combined with pipette blowing, creating a porous, spongy, and spatial structure. The CC FBs produced a variety of signaling factors, showed multidirectional differentiation potential, and grew in a 3D grid in Matrigel, which is necessary for CC-like tissue to maintain a porous structure as a cell scaffold. Within the CC-like tissue, ECs covered the surface of the lumen, and SMCs were located inside the trabeculae, similar to the structure of the primary CC. Various cell components remained stable for 3 days in vitro, but the EC content decreased on the 7th day. Wingless/integrated (WNT) signaling activation led to lumen atrophy and increased tissue fibrosis in CC-like tissue, inducing the same changes in characteristics as in the primary CC. This study describes a preparation method for human artificial CC-like tissue that may provide an improved experimental platform for exploring the function and structure of the CC and conducting drug screening for erectile dysfunction therapy.

NAPG mutation in family members with hereditary hemorrhagic telangiectasia in China.

BACKGROUND: Hereditary hemorrhagic telangiectasia (HHT) is a disease characterized by arteriovenous malformations in the skin and mucous membranes. We enrolled a large pedigree comprising 32 living members, and screened for mutations responsible for HHT. METHODS: We performed whole-exome sequencing to identify novel mutations in the pedigree after excluding three previously reported HHT-related genes using Sanger sequencing. We then performed in silico functional analysis of candidate mutations that were obtained using a variant filtering strategy to identify mutations responsible for HHT. RESULTS: After screening the HHT-related genes, activin A receptor-like type 1 (ACVRL1), endoglin (ENG), and SMAD family member 4 (SMAD4), we did not detect any co-segregated mutations in this pedigree. Whole-exome sequencing analysis of 7 members and Sanger sequencing analysis of 16 additional members identified a mutation (c.784A > G) in the NSF attachment protein gamma (NAPG) gene that co-segregated with the disease. Functional prediction showed that the mutation was deleterious and might change the conformational stability of the NAPG protein. CONCLUSIONS: NAPG c.784A > G may potentially lead to HHT. These results expand the current understanding of the genetic contributions to HHT pathogenesis.

Correlation of Matrix Metalloproteinases-9 with Microemboli Shedding in Carotid Endarterectomy / 中国医学科学院学报

Objective To investigate the change of serum matrix metalloproteinases-9 (MMP-9) expression before,during,and after carotid endarterectomy (CEA) and to investigate the prognostic role of MMP-9. Methods Forty carotid stenosis patients who underwent CEA in the Department of Vascular Surgery,Peking Union Medical College Hospital from February to September 2012 were enrolled in this study. Based on the findings of transcranial doppler monitoring,patients were divided into embolic-positive group and emboli-negative group. Serum samples were obtained from 40 consecutive patients undergoing CEA before operation (pre-op),before de-clamping,30 minutes after de-clamping,and 12 hours after operation (12-h post-op). MMP-9 expression was quantified using enzyme-linked immunosorbent assay and gelatin zymography. Student's t-test and chi-square test were used to compare the differences between these two groups. Results Of these 40 patients,microemboli were detected in 8 patients. The 12-h post-op MMP-9 level was significantly higher than the pre-op level in the emboli-positive group [(904.27±369.47)ng/ml vs. (333.88±126.32) ng/ml,t=4.132,P=0.001].However,there was no difference between pre-op and 12-h post-op MMP-9 levels in the emboli-negative group [(375.83±194.36) ng/ml vs. (472.74±271.21) ng/ml,t=-1.643,P=0.081]. Gelatin zymography also showed higher MMP-9 activity in the emboli-positive group than in the emboli-negative group. Conclusion Serum MMP-9 concentration remarkably increases 12 hours after CEA in patients with microemboli shedding,suggesting MMP-9 may be a potential biomarker for CEA-related cerebral ischemic injury.

Effect of Adequate Anticoagulantion Therapy on the Outcome of Spontaneous Isolated Dissection of Superior Mesenteric Artery / 中国医学科学院学报

Objective To investigate the optimal treatment strategy of spontaneous isolated dissection of superior mesenteric artery (SIDSMA) and the effect of anticoagulation therapy on the prognosis of SIDSMA. Methods The clinical data of 29 patients presented with acute or subacute mesenteric ischemia (a history of less than 14 days) due to SIDSMA admitted to the Department of Vascular Surgery of Peking Union Medical College Hospital from January 1st 2003 to December 31th 2016 were retrospectively analyzed. Results In this study,28 cases were male and the remaining one was female,with an average age of (49.1±7.6) years. The emergency endovascular treatment were performed on 4 cases with severe mesenteric intestinal ischemia,and the symptoms were relieved postoperatively. The remaining 25 cases were treated with conservative treatment. Among 13 cases who were received adequate anticoagulantion therapy,symptoms were relieved or disappeared in 9 cases (69.2%),whereas conservative treatment was ineffective in 4 cases (30.8%),for whom surgical intervention were performed. Among 12 cases who received conservative treatment without sufficient anticoagulation,the abdominal pain was relieved in only 2 cases (16.7%) and the remaining 10 cases (83.3%) were converted to surgical intervention. The success rate of conservative treatment for patients with adequate anticoagulant therapy was significantly higher than that of patients who had not received adequate anticoagulant therapy (P=0.015). Conclusions Adequate anticoagulation therapy has good therapeutic effect in most SIDSMA cases with acute or subacute mesenteric ischemia. For patients with severe mesenteric ischemia or those fail to respond to initial conservative treatment,endovascular treatment may be a more reasonable option.

Research Progress of the Differentiation of 3T3-L1 Preadipocytes into Mature Adipocytes / 中国医学科学院学报

Along with the increase in the prevalence of lipid metabolism disorders,in vitro research on the adipocytes has been a hot topic in recent years. 3T3-L1 preadipocyte line is one of the most commonly used cell line for establishing adipocytes models. However,for 3T3-L1 cell lines,the traditional method,known also as the "Cocktail" method,have disadvantages including low transformation rate and heterogeneous conversion. Therefore,many studies aimed to improving the induction method by changing the time of inducers or adding other new drugs have been performed(known as the improved method). The present study was to summarize the progress of the traditional methods and the improved methods for inducing 3T3-L1 cell lines to mature adipocytes.

Impact of the Action Time of Inducers on Differentiation of 3T3-L1 Cells / 中国医学科学院学报

Objective To explore the effect of the action time of inducers on the differentiation of 3T3-L1 cells to adipocytes. Methods According to the "Cocktail" method,3T3-L1 cells were divided into three groups according to the action time of inducers,with the action time being 2,3 or 4 days,respectively. Cell morphology was observed using inverted microscope and adipose content were detected by Oil red "O" staining and detection of triglyceride. The cell viability was identified by trypan blue staining method. Results The proportion of samples (n=12) with differentiation rate above 80% in group A was 66% (12/18),while the differentiation rate of all the samples (n=18)in group B and group C were above 80%. For the Oil red "O",the OD value at 510 nm in group C was 2.59±0.17,which was significantly higher than that in group A (2.12±0.47;F=6.62,P=0.0001)and group B (2.20±0.17;F=5.15,P=0.0001),while no significant difference was found between group A and group B (F=1.14,P=0.74). As for the triglyceride,the value in group C was (1351.04±119.01)ng/ml,which was significantly higher than that in group A[ (1077.88±272.75)ng/ml;F=6.73,P=0.001] and group B [(1089.38±115.39)ng/ml;F=5.78,P=0.001],while no difference was found between group A and group B (F=0.27,P=0.64). The cell viability in group A,B,and C was (98.3±1.2)%,(98.5±1.8)%,and (98.9±2.1)%,respectively,showing no significant difference (F=0.18,P=0.83). Conclusions The modified procedure for the differentiation of 3T3-L1 cells to adipocytes can increase the differentiation rate and thus may be applied for establishing adipocyte models. The recommended action time is three days.

Advances in the biochemical markers of complications associated with carotid endarterectomy / 中国医学科学院学报

Carotid endarterectomy(CEA)has been proved to be an effective surgery to treat the cerebral ischemia caused by carotid atherosclerotic stenosis. However,there is still no effective mean for the early diagnosis of the CEA-related severe complications such as stroke and death. Many studies have explored the potential biomarkers for stroke alert,although there is still a long way to go for their actual application in clinical settings. The carotid atherosclerotic stenosis,the perioperative complications of CEA,and the stroke share similar pathogenic mechanisms,and some biomarkers such as S100B,matrix metalloproteinase 9,asymmetric dimethylarginine,and neuron-specific enolase have been studied in the clinical trails of CEA. This article summarizes recent advances in this field.

Expressions of p-c-jun and cyclinD1 between vascular smooth muscle cells and endothelial cells exposured to cigarette smoke extract / 中国医学科学院学报

<p><b>OBJECTIVE</b>To investigate the cell viabilities of vascular smooth muscle cells and vascular endothelial cells stimulated by cigarette smoke extract(CSE) .</p><p><b>METHODS</b>The CSE was prepared by smoke-bubbled phosphate buffered saline(PBS) generation.After culturing cells with different concentrations of CSE, we used the cell counting kit-8 to determine the cell viability.The expression levels of c-jun and cyclinD1 were analyzed through Western blot.The c-jun plasmid was transfected to detect the change of cyclinD1 expression.</p><p><b>RESULTS</b>The smooth muscle cell viability increased when the CSE concentration ranged 0.625%-10%, whereas the endothelial cells viability decreased when exposed to the CSE concentration. After exposure to CSE for 48 hours, there was no difference in c-jun expression between toxin group and PBS group;however, the expression of p-c-jun in the smooth muscle cells significantly increased in the toxin groups than in the PBS group(P<0.05) and the expression of p-c-jun in the vascular endothelial cells significantly decreased(P<0.05) . The level of cyclinD1 significantly increased after exposed to CSE, and its expression level also increased in respond to the c-jun overexpression.</p><p><b>CONCLUSION</b>CSE can enhance the proliferation of vascular smooth muscle cells and decrease in the activity of endothelial cells proliferation, which may be explained by the phosphorylation of c-jun and the expression of cyclinD1.</p>

[Study on antiinflammatory effect of a compound TCM agent containing ant extractive in animal models].

OBJECTIVE: To study the antiinflammatory effect of a compound TCM (Traditional Chinese Medicine) agent on animal models. The agent contains ant extractive and a blent of three herbal products, herba epimedii, fructus cnidii, and fructus lycii. METHOD: Three animal models to induce experimental inflammation in rats, including carrageenin--induced paw edema, cotton-ball granuloma and adjuvant induced arthritis, were chosen to study the antiinflammatory effect of the TCM agent. RESULT: The TCM agent showed a marked inhibitory effect on edema induced by all three types of inflammation in rats, the inhibitory rate of the TCM agent at the dose of 0.20, 0.40 and 0.80 g.kg-1 in granuloma model bing over 25% at 1 hour post oral administration, and being 23.8%, 22.7%, 39.7% at 6 hour. In addition, the TCM agent also showed a significant preventive as well as therapeutic effect on adjuvant induced arthritis in rats, and improved the pathological changes of the animal joints with the induced arthritis. CONCLUSION: TCM agent has significant antiinflammatory effects on the three above mentioned animal models.