This study examined the construction of eukaryotic expression plasmid of human transforming growth factor-β3 (hTGF-β3) and its inducing effect on the differentiation of precartilaginous stem cells (PSCs) into chondroblasts. hTGF-β3 gene was amplified by using polymerase chain reaction (PCR) and then inserted into the eukaryotic expression plasmid pcDNA3.1 to construct the eukaryotic expression plasmid pcDNA3.1(+)-hTGF-β3. Rat PSCs were isolated and purified by employing an immunomagnetic cell sorting system. pcDNA3.1(+)-hTGF-β3 was transfected into purified PSCs with the use of linear polyamines. The expression of TGF-β3 and cartilage-specific extracellular matrix (ECM) components was detected after transfection by real-time quantitative PCR, ELISA, immunochemistry and Western blotting, respectively. The results showed that the eukaryotic expression plasmid pcDNA3.1(+)-hTGF-β3 was successfully established as identified by enzyme digestion and DNA sequencing. Real-time quantitative PCR and ELISA revealed that hTGF-β3 was strongly expressed in pcDNA3.1(+)-hTGF-β3-transfected PSCs. Real-time quantitative PCR, immunochemistry and Western blotting showed that the cartilage-specific ECM markers, i.e., cartilage oligomeric matrix protein (COMP), Aggrecan, collagen type X and II were intensely expressed in the pcDNA3.1(+)-hTGF-β3-transfected cells. It was concluded that hTGF-β3 could be stably expressed in pcDNA3.1(+)-hTGF-β3-transfected PSCs and induce the differentiation of PSCs into chondroblasts.

Objective To investigate the possibility of transforming growth factor β (TGF-p) inducing chondrogenesis of precartilaginous stem cells (PSCs) and discuss expression mechanism of extracel-luar matrix. Methods PSCs were induced into a chondrogenic pathway in alginate bead culture in the absence of serum and in the presence of TGF-β1, β2, or-β3. The temporal pattern of expression of cartilage-specific extracellular matrix during chondrogenesis were analyzed by immunocytochemistry, immunoflu-orescence, RT-PCR, immunoprecipitation, Western blot and spectrophotometer. Results Type Ⅱ collagen staining was positive at days 7, 14 and 21 in alginate bead culture, showing most intense staining in the TGF-p3-treated culture. Expression of type Ⅱ collagen was increased in TGF-β3 group. Immunocytochemi-cal analysis of a number of other extracellular matrix components showed widespread expressions of aggre-can, fibromodulin and COMP in alginate bead culture that presented TGF-p3 for 21 days. The expressions of Aggrcan, fibromodulin, type Ⅰ and ⅹ collagen, and COMP were detected by RT-PCR in TGF-β3 group within 8 days, while type Ⅱ collagen began expression at days 8-21. COMP or type X collagen was present in TGF-β3 group at days 7, 14 and 21 by immunoprecipitation or Western blot analysis respectively. The extracted glycosaminoglycan content or the glycosaminoglycan/DNA rate in TGF-βl group was significantly lower than those in TGF-β2 group or TGF-β3 group at days 14 and 21 (P <0.01). Conclusions TGF-β can evocate chondrogenesis of PSCs, when rapid deposition of cartilage-specific extracellular matrix is involved. The sequential events in this pathway leading from the undifferentiated stem cells to mature chon-drocytes can be investigated by analysis of key matrix elements.

Objective To explore the significance of using cytologic and RT-PCR methods to examine(peritoneal) washings and peritoneal tissues of gastric cancer patients in prediction of peritoneal micrometastasis.Methods The peritoneal washings of 38 patients with gastric cancer and 5 patients with benign gastric(lesions) were collected and,at the same time,a small amount of omentum and peritoneum were removed for control.CEAmRNA expression of free cells in peritoneal washings were detected by RT-PCR method and(also) cytology of the washings were performed.Results The CEAmRNA expression rate of peritoneal washings and peritoneal tissues were 36.8%(14/38) and 39.5%(15/38)respectively.Both were more(sensitive) than that of cytologic examination 26.3%(10/38).TNM staging,depth of invasion,lymph node metastasis,and serosal involvement were related to the expression rate of CEAmRNA.Conclusions mRNA of CEA is more sensitive and specific than cytologic examination for detecting free cancer cells in peritoneal cavity.It is an effective method for detecting peritoneal micrometastases in gastric cancer patient.

This study examined the construction of eukaryotic expression plasmid of human transforming growth factor-β3 (hTGF-β3) and its inducing effect on the differentiation of precartilaginous stem cells (PSCs) into chondroblasts.hTGF-β3 gene was amplified by using polymerase chain reaction (PCR) and then inserted into the eukaryotic expression plasmid pcDNA3.1 to construct the eukaryotic expression plasmid pcDNA3.1(+)-hTGF-β3.Rat PSCs were isolated and purified by employing an immunomagnetic cell sorting system.pcDNA3.1(+)-hTGF-β3 was transfected into purified PSCs with the use of linear polyamines.The expression of TGF-β3 and cartilage-specific extracellular matrix (ECM)components was detected after transfection by real-time quantitative PCR,ELISA,immunochemistry and Western blotting,respectively.The results showed that the eukaryotic expression plasmid pcDNA3.1(+)-hTGF-β3 was successfully established as identified by enzyme digestion and DNA sequencing.Real-time quantitative PCR and ELISA revealed that hTGF-β3 was strongly expressed in pcDNA3.1(+)-hTGF-β3-transfected PSCs.Real-time quantitative PCR,immunochemistry and Western blotting showed that the cartilage-specific ECM markers,i.e.,cartilage oligomeric matrix protein (COMP),Aggrecan,collagen type Ⅹ and Ⅱ were intensely expressed in the pcDNA3.1(+)-hTGF-β3-transfected cells.It was concluded that hTGF-β3 could be stably expressed in pcDNA3.1(+)-hTGF-β3-transfected PSCs and induce the differentiation of PSCs into chondroblasts.

Fibrotic diseases often cause damage to multiple tissues and organs, which still lack an effective therapy. At present, it has been found that long non-coding RNA(LncRNA) H19 is differentially expressed in fibrosis of skin, liver, lung, heart, kidney, and other organs, indicating that LncRNA H19 may be involved in the occurrence and development of fibrotic diseases.This article summarizes the relationship between LncRNA H19 and the development of fibrotic diseases and its role in various organs, which provides a new idea for the diagnosis, prevention, and treatment of fibrotic diseases.

Fibrotic diseases often cause damage to multiple tissues and organs, which still lack an effective therapy. At present, it has been found that long non-coding RNA(LncRNA) H19 is differentially expressed in fibrosis of skin, liver, lung, heart, kidney, and other organs, indicating that LncRNA H19 may be involved in the occurrence and development of fibrotic diseases.This article summarizes the relationship between LncRNA H19 and the development of fibrotic diseases and its role in various organs, which provides a new idea for the diagnosis, prevention, and treatment of fibrotic diseases.

ObjectiveTo observe the clinical efficacy and possible mechanisms of Xianglian Huazhuo Granules （香连化浊颗粒， XHG） in the treatment of chronic atrophic gastritis with intestinal metaplasia. MethodsA total of 140 patients with chronic atrophic gastritis and intestinal metaplasia were randomly divided into a treatment group and a control group， with 70 cases in each group. The treatment group received 12.5 g of XHG orally， twice daily. The control group received 12.5 g of placebo orally， twice daily. Both groups were treated for 6 months. The traditional Chinese medicine （TCM） symptom scores， pathological types， serum tumor markers of the digestive system， and serum bile acids （TBA）， interleukin-23 （IL-23）， and Dickkopf-related protein 1 （DKK-1） levels were observed before and after treatment. Safety indicators and adverse events were recorded. After treatment， TCM syndrome efficacy and pathological types were evaluated， and patients were followed up for 18 months with gastric endoscopy and pathological results， which were compared with the results after treatment finished. ResultsTwo patients dropped out in the control group， and a total of 168 cases were included in the final analysis， 70 in the treatment group and 68 in the control group. The treatment group showed a significant reduction in TCM symptom scores， serum TBA， IL-23， and DKK-1 levels， and a significant increase in alpha-fetoprotein （AFP）， carbohydrate antigen 125 （CA125）， carbohydrate antigen 199 （CA199） levels； in the control group， carcinoembryonic antigen （CEA）， CA125， CA199 levels significantly increased （P＜0.05 or P＜0.01）； and carbohydrate antigen 242 （CA242） level in both the treatment group and the control group decreased significantly （P＜0.01）. The treatment group had lower TCM symptom scores and lower levels of serum TBA， IL-23， and DKK-1 compared to the control group （P＜0.05）. The effective rate for TCM syndrome efficacy in the treatment group was 80.00% （56/70）， significantly higher than the 20.59% （14/68） in the control group （P < 0.05）. The effective rate for pathological classification in the treatment group was 72.73% （8/11） for mixed intestinal metaplasia， significantly better than 46.15% （6/13） in the control group （P＜0.05）. No adverse events were reported in either group. Among 40 patients who had a follow-up endoscopy after one year， 21 were from the treatment group， of whom 11 showed reduced intestinal metaplasia， 9 showed no significant changes， and 1 had worsened； while 19 patients in the control group had 4 with reduced intestinal metaplasia， 13 with no significant changes， and 2 with worsened conditions. No cancer was detected in either group. The treatment group showed significantly better improvement in intestinal metaplasia on follow-up gastric endoscopy pathology than the control group （P＜0.05）. ConclusionXHG can significantly improve the clinical symptoms in patients with chronic atrophic gastritis and intestinal metaplasia and reduce the degree of mixed intestinal metaplasia. The mechanism may involve lowering serum TBA， DKK-1， and IL-23 levles， thus delaying the progression from inflammation to cancer.

Objective: To compare the clinical effect of the Yu flap and the Karapandzic flap in repairing greater than 2/3 defects of the lower lip and to provide a reference for clinical application. Methods: Ten patients with greater than 2/3 lower lip defects after surgical resection of lower lip tumors and vascular malformations were enrolled: 5 patients were repaired with the Yu flap (Yu flap group) and 5 patients were repaired with the Karapandzic flap (Karapandzic flap group). Follow-up for at least 1 year was conducted to evaluate the morphology (symmetry, stoma, exposure of vermilion) and function (sensory function, motor function) of the reconstructed lower lip. Results : All the flaps survived, and all wounds showed primary healing. The lower lips reconstructed with the Yu flap or the Karapandzic flap obtained similar satisfactory oral function. The sensory function was essentially restored. There were no obvious obstacles in speech and expression, and no saliva leakage occurred. In the Yu flap group, only 1 patient had slight microstomia. In the Karapandzic flap group, 2 patients had slight microstomia and 3 patients had moderate microstomia. 90% (9/10) of the patients were very satisfied with the postoperative outcome, and 1 patient in the Karapandzic flap group was basically satisfied. Conclusion Both the Yu flap and the Karapandzic flap can be used to repair greater than 2/3 lower lip defects and reliable outcomes can be achieved. These two methods can achieve similar oral functions, but the effect of the Karapandzic flap is inferior to that of the Yu flap in terms of aesthetic appearance, and microstomia often occurs, while the Yu flap can generally maintain the original size of the mouth cleft.