To clarify the function and molecular mechanism of miR-155 in myogenic differentiation of C2C12, we constructed adenovirus over-expression vector of miR-155, then C2C12 cells were infected by adenovirus and induced myogenic differentiation. First, we observed the morphology of C2C12 after differentiation. Then the mRNA and protein expressions of myogenic markers (MyoD, MyoG and MyHC) were detected by qPCR and western blotting. Subsequently, the dual luciferase reporter gene assay was carried out to validate putative target gene (TCF4) of miR-155. Meanwhile, mRNA level of TCF4 was analyzed after over-expressing miR-155. The results show that over-expressed miR-155 reduced myotubes formation. Moreover, the mRNA and protein expression of MyoG and MyHC decreased significantly (P < 0.01). Further research demonstrated miR-155 bound the one (4532-4538) of three putative sites (1487-1493,1516-1522, 4532-4583) of TCF4 by luciferase reporter gene assay and the mRNA level of TCF4 decreased notably (P < 0.05). The data suggest that miR-155 inhibited myogenic differentiation of C2C12 through targeted TCF4.
Animals ; Basic Helix-Loop-Helix Leucine Zipper Transcription Factors ; genetics ; Cell Differentiation ; Cell Line ; Genetic Vectors ; Mice ; MicroRNAs ; genetics ; Myoblasts ; cytology ; Myogenin ; genetics ; metabolism ; Myosin Heavy Chains ; genetics ; metabolism ; RNA, Messenger ; genetics ; Transcription Factor 4

To explore the plasticity of human amniotic mesenchymal cells(hAMCs) into cardiomyocyte-like cells,hAMCs were isolated from human amnion with collagenase digestion.Phenotype of the isolated cells was analyzed by flow cytometry(FCM).hAMCs were treated with 5-azacytidine and basic fibroblast growth factor(bFGF) to investigate their ability of differentiation into cardiomyocytes.The induced differentiated cells were evaluated by immunofluorescence for desmin and ?-actin expression and by RT-PCR for Nkx2.5,GATA-4 and alpha-myosin heavy chain(?-MHC) mRNA expression.The results showed that,after primary culture,hAMCs could reach a confluence of 80% with swirl like growth at 6 days.The cells proliferated rapidly after passages with a 100% confluence at 3~4 d.hAMCs were positive expression of CD44 and vimentin,but negative for CK19.After induced differentiation at 8~10d,the differentiated cells have close-up arranged with long spindle-shape.At 2 weeks and 4 weeks,induced cells expressed ?-actinin and cardiac-specific transcription factor Nkx2.5.Expressions of GATA-4 and desmin can be detected but ?-MHC can not in the hAMCs both before and after the induction.In conclusion,hAMCs have the ability of differentiation into cardiomyocyte-like cells,which means that hAMCs can be regarded as candidate cells for cellular cardiomyoplasty(CCM).

BACKGROUNDSo far, there is no efficient treatment for pulmonary fibrosis. The objective of this study was to determine whether intramuscular injection of the hepatocyte growth factor (HGF) plasmid DNA by in vivo electroporation could prevent bleomycin-induced pulmonary fibrosis in rats, and to investigate the possible mechanisms.

METHODSTwenty male Wistar rats were randomly divided into four groups: control group (group C), model group (group M), early intervention group (group I) and late intervention group (group II). Groups M, I and II were intratracheally infused with bleomycin, then injected the plasmid pcDNA3.1-hHGF to group I on day 7, 14 and 21. Group II received the same treatment like Group I on day 14 and 21. All the rats were killed on day 28 after bleomycin injection. We detected Homo HGF expression in the rats with ELISA method and estimated the pathological fibrosis score of lung tissue using hematoxylin eosin (HE) and Massion staining. The mRNA expression of transforming growth factor-beta1 (TGF-beta1), cycloxygenase-2 (COX-2), and rat HGF in rat pulmonary parenchyma were evaluated by RT-PCR. Immunohistochemistry and Western blotting were performed to determine the protein expression of transforming TGF-beta1 and COX-2 in lung parenchyma.

RESULTSThe plasmid pcDNA3.1-hHGF could express hHGF in NIH3T3 cells and the hHGF protein is secreted into the culture medium. The expression of hHGF protein could be monitored in quadriceps muscle, plasma and lung in Groups I and II. Pulmonary fibrosis levels of Groups I and II were obviously lower than that of group M (P < 0.05). Expression of TGF-beta1 protein and mRNA in lung tissue was markedly decreased in Groups I and II compared with Group M (P < 0.05). The level of expression of HGF and COX-2 mRNA was higher in Groups I and II than in Group M (P < 0.05).

CONCLUSIONSInjection of the plasmid pcDNA3.1-hHGF into skeletal muscle with electroporation has a potential role in the treatment of bleomycin-induced lung fibrosis. Exogenous HGF may inhibit the expression of TGF-beta1 and regulate the crosstalk between AECs and mesenchymal fibroblasts.

Animals ; Bleomycin ; toxicity ; Cyclooxygenase 2 ; genetics ; Electroporation ; Genetic Therapy ; Hepatocyte Growth Factor ; genetics ; Hydroxyproline ; analysis ; Injections, Intramuscular ; Lung ; chemistry ; Male ; Muscle, Skeletal ; metabolism ; Plasmids ; Pulmonary Fibrosis ; chemically induced ; therapy ; RNA, Messenger ; analysis ; Rats ; Rats, Wistar ; Transforming Growth Factor beta1 ; analysis ; genetics

This study aims to construct a dynamic two-dimensional characterization technique for the hygroscopicity of traditional Chinese medicine extracts and investigate the effect of material properties of powders on hygroscopicity. The dynamic hygroscopicity-time curves of the powders were measured at 25 ℃ and 75% humidity, and the semi-equilibrium hygroscopicity time (t_1/2) and equilibrium hygroscopicity (F^∞) were derived as two-dimensional evaluation indicators. Finally, the correlation between the material properties and the hygroscopic behavior was analyzed by principal component analysis (PCA) and partial least squares analysis (PLS). The results showed that the dynamic two-dimensional characterization system of hygroscopicity constructed with 1/t_1/2 = 0.1 h^-1 and F^∞ = 15% as the center can classify the hygroscopic behavior of traditional Chinese medicine extracts into four categories: fast hygroscopicity with strong hygroscopicity, slow hygroscopicity with strong hygroscopicity, fast hygroscopicity with weak hygroscopicity and slow hygroscopicity with weak hygroscopicity. The moisture absorption was negatively correlated with D₅₀, D₉₀, ρ_b and ρ_t; the moisture absorption rate was negatively correlated with D₁₀, D₅₀, D₉₀, ρ_b, ρ_t, and positively correlated with moisture content. The hygroscopicity dynamic two-dimensional characterization indicators of Chinese medicine extracts (CMEs) constructed in this study matched with the physical properties. The method of dynamic multi-dimensional characterization technology is feasible and scientific, and the idea has strong promotional value.

Objective To investigate the effect of intravenous fluid infusion combined with water throuth gastrointestinal tract in treating the patients with hyperosmolar hyperglycemic state(HHS).Methods 30 HHS pa- tients were recruited.All the patients were given water throuth gastrointestinal infusion while they were administrat- ed continuously intravenous infusion.Laboratory parameters such as serum natrium,serum potasium,serum glucose and serum osmolarity ect were monitored at the admission and after treatment.Results Serum glucose and serum osmolarity of HHS patients were decreased smoothly at the speed of less than 3 mmol?L~(-1).h~(-1)during the first 12 hours after treatment.After 48 hour-treatment,serum natrium,serum potassium and serum osmolarity recovered to normal levels except 2 deaths,serum glucose decreased to(10.8?5.2)mmol/L.Conclusion Intravenous fluid infu- sion combined with water throuth gastrointestinal tract for the patients could lower smoothly serum glucose and serum osmolarity and decrease the mortality of the HHS patients.

Objective To study the change and rule of immunological function among the patients with coal arsenic poisoning in order to provide a basis for tumor risk evaluation and monitoring.Methods Seventy patients with coal arsenic poisoning aged from 24 to 71 years old(44 men,26 women,averaging 41 years old)were divided into 4 groups including 23 cases having a course less than 10 years,21 case8 lasting for 10～19 years,20 cages for more than 20 years,6 cases of cancer,and 26 healthy normal controls.Flow cytometer(FCM)was used to analyze the frequency of CD3+(total T cell),CD3+CD4+(inducer/helper T cell),CD3+CD8+(suppressor/cytotoxic T cell),CD19+(B lymphocyte),and CD56+CD16+(natural killer cell)lymphocyte subsets in the peripheral blood of the subjects and the expression rates of lymphocytic membrane surface molecules of human leucocvte antigen (HLA)-DR,CD25,CD38 were also determined by FCM.Results The pmportions of CD3+cells in periDheral blood of less than 10 years,10～19 years,more than 20 years and cancer groups were (63.76±9.32)%。(55.63± 12.97)%,(51.00±12.23)%and(49.83±,9.89)%respectively,which were significantly lower than that in control group[(68.10±8.62)%],and there was a significant difference between different groups(F=12.862,P<0.05). In less than 10 years,10～19 years,more than 20 years and cancer groups,the proportion of CD3+CD4+cells cells was (31.35±6.62)%,(28.38±8,66)%,(24.13±6.46)%and(19.17±4.96)%respectively,which wag significantly lowerthan that in control group[(34.28±7.32)%],and significant in a-group difference was found(F=10.455, P<0.05).The percentages of CD19+cells in more than 20 yeats and cancer groups[(9.00±5.32)%,(9.00± 3.29)%]were lower than that in control group and less than 10 years group[(11.80±3.43)%,(12.35±4.53)%] (P<0.05),while no statistical difference was found between other groups.The expression rates of CD25 and CD38 in lymphocytes of cancer group[(17.96 ±4.98)%,(41.38±8.54)%]were obviously higher than those in control group[(13.10±338)%,(28.60±5.51)%]and there were statistical differences between the experimental groups(P<0.05).The expression rate of HLA-DR in 10～19 years groups[(18.20±6.25)%]was significantly higher than that in control group[(10.72±7.06)%]and less than 10 years group[(11.78±5.13)%],while it was the same in more than 20 years and cancer group[(20.30±8.01)%,(21.82±10.97)%].Conclusions Reduction of cellular immune function caused by coal arsenic poisoning may be an important mechanism of skin cancer.CelMar immune function may be used as a warning signal of skin cancerization of patients with coal arsenic poisoning.

Objective To improve the diagnosis accuracy and treatment quality of juxtaglomerular cell tumor.Methods The clinical data of 2 female patients(20 and 36 years,respectively)with juxtaglo- merular cell tumor were presented and discussed in combination with review of the literature,including the onset characteristics,imaging features,treatment,pathology and prognosis.Case 1 presented with hyperten- sion of 180/110 mm Hg.The laboratory examinations showed that in decubitus and standing position,the plasma rennin activity(PRA)was 3.2?g - L~(-1)?h~(-1)and 36.5?g?L~(-1)?h~(-1);angiotensinⅡwas 54.3 pg/ml and 183.5 pg/ml;aldosterone was 193.5 prnol/L and 489.4 pmol/L,respectively;serum kalium was 2.6 mmol/L.Case 2 presented with hypertension of 210/120 mm Hg.The laboratory examination results were as follows:in decubitus and standing position,PRA was 4.3?g?L~(-1)?h~(-1)and 37.0?g?L~(-1)?h~(-1);angio- teusinⅡwas 55.6 pg/ml and 200.4 pg/ml;aldosterone was 162.4 pmol/L and 506.3 pmol/L,respectively; serum kalium was 3.0 mmol/L.On CT scan,both cases had renal tumor,with the diameter of 3.0 cm and 3. 5 cm,respectively.Results Case 1 underwent laparoscopic partial nephrectomy.Case 2 who had artery stricture and severe functional injure of the right kidney underwent laparoscopic right nephrectomy.The oper- ative time was 3.0 h and 2.0 h,and the blood loss was 175 ml and 112 ml,respectively.There was no mor- tality or postoperative complication.In 1 or 2 postoperative weeks,Case 1 had blood pressure(BP)of 120/70 mm Hg;in deeubitus and standing position,PRA was 1.5?g ~ L~(-1)?b~(-1)and 12.8?g?L~(-1)?h~(-1);angio- tensinⅡwas 30.6 pg/ml and 97.5 pg/ml;aldosterone was 78.5 pmol/L and 192.2 pmol/L,respectively ;se- rum kalium was 4.2 mmol/L.Case 2 had BP of 125/75 mm Hg;in decubitus and standing position,PRA was 1.6?g.L~(-1)?h~(-1)and 12.3?g.L~(-1)?h(-1);angiotensinⅡwas 34.3 pg/ml and 83.5 pg/ml;aldoste- rone was 62.6 pmol/L and 292.5 pmol/L,respectively;serum kalium was 4.8 mmol/L.Pathology showed that the juxtaglomerular cell tumor had intact envelop.Light microscopically,the tumor was very much like a hemangiopericytoma,showing active proliferation and nuclear atypia.The immunohistochemical staining showed positive Vimentin,CD_(34)expression,and negative MSA,EMA,Bcl-2,?-SmA,AG/AG3,34?En, CD_(117),CD_(31),Iv glue,Ki-G~-(＜2%)expression.Ultrastructural changes of the nuclei and some organelles in the cytoplasm were observed under electron microscope.The conspicuous ultrastructural feature was the pres- ence of secretion granules and rhomboid-shaped,crystal-like structures in the dilated cisternae of rough endo- plasmic reticulum and vesicles of Golgi complex.The follow-up was 14 and 6 months,respectively;the renal function was normal and no tumor recurrence was found.Conelusions Juxtaglomerular cell tumor is a rare tumor which can produce renin.It is characterized by severe hypertension and low serum potassium.La- boratory examination results are helpful for the diagnosis:PRA and angiotensinⅡincrease obviously ;aldoste- rone is 1-10 times more than normal;serum kalium is commonly between 2.1-3.5 mmol/L.The definite diagnosis depends on clinical presentations,immunohistochemistry,light and electron microscopic examina- tions.Laparoscopie operation is the first choice of surgical treatment.

At present, the modernization of Chinese medicine preparations (CMPs) is still a challenging task. The 3 typical Chinese medicine materials (CMMs) used for preparing CMPs are the powders, extracts, and components of Chinese medicine and their properties of CMMs are important for designing CMPs. Basing on our long term research, we have established a property system for CMMs according to the state of CMMs under an exactly condition and according to the interaction characteristics between substances. The property system could be divided into 5 categories: material composition, spatial structure, body property, surface property, physicochemical properties, and they could also be divided into 18 subcategories. Furthermore, we also established the corresponding index and characterization system, where the 61 indexes and characterization techniques were systematically summarized. At last, we hope that the article will promote the modernization of CMPs.

Objective:To explore the effects of different approaches for second-trimester multifetal pregnancy reduction on pregnancy outcome in women with dichorionic triamniotic (DCTA) triplet.Methods:A retrospective study was performed on 51 women with DCTA triplet pregnancies who were referred to Guangdong Women and Children Hospital for second-trimester multifetal pregnancy reduction from January 2014 to January 2020. All participants were divided into either preventive group ( n=39) or treatment group ( n=12) according to the indication for multifetal pregnancy reduction, and they were further allocated to three subgroups based on different reduction methods, which were reduction to dichorionic twin by radiofrequency ablation (RFA) (RFA subgroup), reduction to monochorionic singleton (KCl-singleton subgroup) or monochorionic twin (KCl-twin subgroup) by cardiac injection of potassium chloride. Pregnancy loss rate, neonatal birth weight, gestational age at delivery, incidence of intrauterine death, and neonatal death were compared and analyzed between different groups using t-test, analysis of variance, Chi-square test, Fisher's exact test and Bonferroni correction. Results:(1) The mean gestational week at operation in the treatment group was significantly later than that in the preventive group [(18.5±3.1) vs (15.0±2.3) weeks, t=-4.209, P<0.001]. In the preventive group, the mean gestational week at operation in the RFA subgroup was later than the KCl-singleton and KCl-twin subgroup[(17.2±1.6) vs (13.8±1.5) and (12.7±1.0) weeks, t=6.630 and 3.875, respectively, both P<0.05]. (2) The postoperative pregnancy loss rate in the preventive group was decreased compared with the treatment group [10.3%(4/39) vs 5/12, Fisher's exact test, P<0.05], and the live birth ratio was increased [ 85.7%(48/56) vs 10/18, χ2=5.640, P=0.018]. No live birth infants with birth weight <1 500 g was reported in the KCl-singleton subgroup in preventive group, and the statistical significance was observed in the intra-group differences ( P<0.05) rather than the pairwise comparison differences in the preventive group. For the proportion of live births, there was a statistically significant difference in the intra-group comparison in the treatment group, which was higher in the RFA subgroup than that in the KCl-twin subgroup (6/6 vs 1/6, P=0.045). No significant difference was revealed among pregnancy loss rate, gestational weeks at delivery, the mean birth weight, premature delivery <32 gestational weeks, and full-term birth rate among three different approaches within the two groups. (3) No monochorionic twin complications or perinatal death occurred in any RFA or KCl-singleton subgroups in the two groups. In the KCl-twin subgroups including five cases with ten fetuses, including three live birth, four miscarriage, three intrauterine death occured, while no neonatal death was reported. One case with selective fetal uterine growth restriction in the preventive group delivered two live births, and one case with twin-to-twin transfusion syndrome in the treatment group had intrauterine death in one fetus and one survival neonate. Conclusions:The pregnancy outcome of multifetal pregnancy reduction to dichorionic diamniotic twins by RFA or reduction to singleton by cardiac injection of potassium chloride are comparative in women with DCTA triplet, regardless of whether it is a preventive or therapeutic reduction.

OBJECTIVETo investigate the reconstruction of wide vermilion and orbicularis oris muscle defect with satisfactory outcome of aesthetics, sensation, and function.

METHODSThe buccal musculomucosal flap based on the anterior buccal branches of the facial artery was used to reconstruct wide defect of vermilion and orbicularis oris muscle on upper or lower lip.

RESULTS7 patients were treated. 5 cases had no postoperative complication. Partial mucosal necrosis on the tip of the flaps happened in 2 cases, but the underlying muscle survived and was re-mucosalized spontaneously. No other complication was observed. The sensation of cold, heat and touch could be detected on the first postoperative day. Electromyographic and electron microscopic studies confirmed innervation of the muscle in the flap.

CONCLUSIONSThe buccal musculomucosal flap is a reliable reconstruction option for wide defect of vermilion and orbicularis oris muscle which can' t be reconstructed with conventional method. Satisfactory aesthetic and functional results can be achieved with the buccal musculomucosal flap.

Adolescent ; Adult ; Cheek ; Facial Muscles ; pathology ; transplantation ; Female ; Follow-Up Studies ; Humans ; Lip ; pathology ; Male ; Middle Aged ; Mouth Mucosa ; transplantation ; Mouth Neoplasms ; pathology ; surgery ; Neoplasm Staging ; Reconstructive Surgical Procedures ; methods ; Surgical Flaps ; Young Adult