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C-fos is a transcription factor that plays an important role in cell proliferation, differentiation and tumor formation. The aim of this study was to clone the goat c-fos gene, clarify its biological characteristics, and further reveal its regulatory role in the differentiation of goat subcutaneous adipocytes. We cloned the c-fos gene from subcutaneous adipose tissue of Jianzhou big-eared goats by reverse transcription-polymerase chain reaction (RT-PCR) and analyzed its biological characteristics. Using real-time quantitative PCR (qPCR), we detected the expression of c-fos gene in the heart, liver, spleen, lung, kidney, subcutaneous fat, longissimus dorsi and subcutaneous adipocytes of goat upon induced differentiation for 0 h to 120 h. The goat overexpression vector pEGFP-c-fos was constructed and transfected into the subcutaneous preadipocytes for induced differentiation. The morphological changes of lipid droplet accumulation were observed by oil red O staining and bodipy staining. Furthermore, qPCR was used to test the relative mRNA level of the c-fos overexpression on adipogenic differentiation marker genes. The results showed that the cloned goat c-fos gene was 1 477 bp in length, in which the coding sequence was 1 143 bp, encoding a protein of 380 amino acids. Protein structure analysis showed that goat FOS protein has a basic leucine zipper structure, and subcellular localization prediction suggested that it was mainly distributed in the nucleus. The relative expression level of c-fos was higher in the subcutaneous adipose tissue of goats (P < 0.05), and the expression level of c-fos was significantly increased upon induced differentiation of subcutaneous preadipocyte for 48 h (P < 0.01). Overexpression of c-fos significantly inhibited the lipid droplets formation in goat subcutaneous adipocytes, significantly decreased the relative expression levels of the AP2 and C/EBPβ lipogenic marker genes (P < 0.01). Moreover, AP2 and C/EBPβ promoter are predicted to have multiple binding sites. In conclusion, the results indicated that c-fos gene was a negative regulatory factor of subcutaneous adipocyte differentiation in goats, and it might regulate the expression of AP2 and C/EBPβ gene expression.
Subject(s)
Animals , Goats/genetics , Cell Differentiation/genetics , Adipogenesis/genetics , Gene Expression Regulation , Proteins/genetics , Cloning, MolecularABSTRACT
Objective:To analyze the clinical and physical tactors associated with acute bone marrow suppression in concurrent chemo-radiotherapy for rectal cancer and to provide a reference standard for the best clinical treatment plan. Methods:Retrospective analy-sis was performed on 62 patients with rectal cancer who received concurrent radiotherapy and chemotherapy in our department. The pelvis was contoured for each patient in the radiotherapy treatment planning system and divided into three subsites: lumbosacral spine, ilium, and lower pelvis. Prognostic clinical and physical factors were analyzed by univariate and multivariate analyses. Evaluated prognostic clinical factors included sex, age, clinical stage, original hemoglobin levels, and chemotherapy, operation, and radiation modes;physical factors included V5, V10, V15, V20, V25, V30, V35, V40, V45, V50, Dmax, and Dmean of lumbosacral spine, ilium, low-er pelvis, and pelvis. Results:The percentage of patients who developed acute bone marrow suppression (≥2 grade) was 61.3%(38/62).Univariate analysis of related factors revealed statistically significant differences were sex, chemotherapy, lumbosacral spine V45, il-ium V20, and ilium V30. Multivariate logistic regression analysis indicated that chemotherapy and ilium V30 are the risk factors for acute bone marrow suppression. The receiver operating curve showed that the threshold of ilium V30 was 44%. Conclusion:Acute bone marrow suppression is influenced by more than one factor;local control rate of the tumor and acute bone marrow suppression are tradeoffs in rectal cancer treatment. An appropriate chemotherapy method should be selected, and ilium V30 must be maintained below 44%to prevent bone marrow suppression in rectal cancer patients.
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Objective To evaluate the safety,tumor radical and early postoperative efficacy through comparison of laparoscopic gastric D2 radical surgery with traditional open gastric D2 radical surgery.Methods 254 patients with upper stomach cancer underwent surgery were selected,132 cases using conventional open gastrectomy (the traditional laparotomy group),122 patients underwent laparoscopic radical gastrectomy (laparoscopic surgery group).Laparoscopic surgery group with traditional open surgery group had no statistically significant differences in gender,age,tumor location,histological type and TNM staging.Results Open surgery group and laparoscopic surgery group had statistically significant differences in operative time [(235.78±31.56) min,(256.43±54.08) min,P ＜ 0.001],blood loss [(326.69±89.73) ml,(158.31±62.98) ml,P ＜ 0.001],incision length [(16.53±2.34) cm,(5.51±1.15) cm,P ＜ 0.001],gastrointestinal recovery time [(4.22±0.91) d,(3.31±0.83) d,P ＜ 0.001],first time eating liquid [(5.78±0.95) d,(5.56±0.78) d,P ＜ 0.001] and postoperative hospital stay [(12.62±2.89) d,(11.18±1.78) d,P ＜ 0.001].The total number of lymph node dissection and complications was not statistically significant.Conclusions Laparoscopic gastric D2 radical surgery is a safe,minimally invasive surgical method.Laparoscopic gastric D2 radical surgery has the same lymph node dissection and good early outcome compared with the traditional gastric D2 radical surgery,but postoperative recovery fast and less invasive.
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Objective To analyze psychological status of the donors and the recipients before and after the relative kidney transplantation.Method Symptom checklist 90 (SCL-90) were performed for 147 renal transplant recipients and donors preoperative and postoperative.Statistical analysis were performed to analyze the scores between the recipients,donors and normal standards.Results Over 90％ recipients were of obvious anxiety preoperative.The scores of most factors of recipients were significantly higher than those of donors.The scores of somatization,interpersonal sensitivity,depression,anxiety,hostility,phobia and paranoia between two groups has statistical significance(P＜0.05).Part of recipients retest SCL-90 3 month after operation,the data showed that the scores of somatization,interpersonal sensitivity,depression,anxiety,hostility were obviously declined after operation(P＜0.05).And the test also showed that most of the donors were willing to help preoperative.Conclusion Preoperative psychological test was useful in preoperational psychological intervention for transplantation recipients.It can increase the safety of the recipients during perioperative period.
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Objective To establish a stable and efficient method of culturing imDCs in vitro,and to explore the effect of GW5074, which blocks ERK1/2 signal pathway in the process of imnature dentritic cells (imDCs) on inducing differentiation of the na(i)ve allogeneic CD4+ T cells into Treg cells in vitro. Methods The imDCs and mature DCs (mDCs) were isolated and cultured from the peripheral blood mononuclear cells (PBMC) derived from a healthy adult male volunteer, and they were identified by cell morphology, cell surface marker and cell functions respectively. Na(i)ve CD4+ T cells were isolated from newborn umbilical vein blood and were divided into 5 groups to be cultured: (1) Blank control group: Na(i)ve CD4+ T cells were cultured alone;(2) Positive control group: The irrDCs were Middle-concentration GW5074 group;(5) High-concentration GW5074 group. In the last three groups, imDCs and na(i)ve CD4+ T cells were co-cultured, the same as the positive control group, but these groups were added by GW5074 dilution at the concentrations of 8, 24, and 40μmol/Lrespectively. After co-culture for 5 days, the transformation ratio from naive CD4+T cells to Treg T cells was detected by flow cytometry. Results On the surface of imDCs, there was stronger pression of CD1a, but weaker expression of CD80 and CD83. On the contrary, on the surface of mDCs, there was weaker expression of CD1a, but stronger expression of CD80 and CD83. The stimulation index in imDCs group and mDCs group was 1.12±0.03 and 2.85±0. 07 respectively. The transformation ratio of Treg T cells in blank control group, positive control group, low-concentration GW5074 group, middle-concentration GW5074 group and high-concentration GW5074 group was (5. 81±1.36)%, (35.73±2.07)%, (22.53±2.11)%, (11.55±1.73)%, and (4.97±1.83)%respectively. One-way ANOVA analysis revealed that there was no significant difference between high-concentration GW5074 group and blank control group, P＞0. 05, but significant difference between the remaining groups, P＜0.01. Conclusion High purity of imDCs can be obtained from PBMC by induction with rhGM-CSF and rhIL-4. ERK1/2 signal pathway plays a role in inducing the immune tolerance. GW5074 can inhibit differentiation of na(i)ve CD4+ T cells into Treg T cells.
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Objective To investigate the distribution and clonality of TCR V? repertoire T cells in peripheral blood mononuclear cells with chronic myeloid leukemia(CML)at chronic phase(CP)and complete remission(CR).Methods The CDR3 of TCR V? 8 subfamily genes were amplified in mononuclear cells of peripheral blood from CML patients in CP(8 cases)and CR(8 cases)phases for observation of the usage of TCR V? repertoire by RT-PCR.The positive PCR products were further labeled with fluorescence and analyzed by genescan technique for the CDR3 size for evaluation of the clonality of the detectable TCR V? T cells.A total of 10 cases of healthy adults served as controls.Results The mean values of detectable TCR V? subfamilies were(3.25?0.89)in patients with CML-CP,predominantly in V?1,2 and V?3,whereas(3.5?0.76)of 8 V? subfamily T cells were expressed in patients with CML-CR,predominantly in V?1,2,3 and V?8.There was no significant difference between the above groups and the healthy adults(3.5?0.52).In addition,a higher frequency expression of V?8 could be found from peripheral blood in CR phase(5/8)rather than from that in CP phase(2/8)and from that of the healthy adults(3/10).Genescan analysis showed that the majority of PCR products from both CML and healthy adults displayed clonal expansion,predominantly in V?1,V?2 and V?3.Clonal expanded V?7 subfamily T cells,however,could be identified from two patients in CR phase but not in all of 10 healthy adults.Conclusion The similar distribution and clonal pattern of TCR V? repertoire T cells can be found in peripheral blood from the three groups,suggesting that the cellular immunosuppression of CML patients might not be mainly involved in the alteration of TCR V? repertoire.In CML-CR groups,the expression and clonal pattern of some V? repertoires are different from those in other groups,which needs further studies.
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Objective To establish a HPLC method for the determination of geniposide in Mengning Tablets and to investigate the thermal stability of geniposide. Methods The content of geniposide in Mengning Tablets was determined on a E- clipse XDB-C_(18) column with a mobile phase consisting of 0.2% triethylamine (containing 0.1% phosphate) and acetonitrile (85: 15) at a flow rate of 1.0 mL?min~(-1) and detected at a wavelength of 240 nm. The thermal stability of geniposide was investigated by hyperthermal accelerated test. Results A linearity was obtained from 0. 10 ?g to 0.52 ?g of geniposide (r=0.9997, n=5) and the recovery was 98.92%(n=5). The variation of geniposide in Mengning Tablets was in first-order reaction and the expiry date of the medicine at normal temperature was 2.83 years when geniposide content was used as criteria. Conclusion This is a fast method for the determination of the expiry date of a Chinese patent medicine and this method is useful infor the development of a new medicine.
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AIM: To establish the quality standard of Puqinxiaoyan Tablets (Radix Scutellariae, Herba Taraxaci, Herba Corydalis Bungeanae, etc.). METHODS: TLC was used for identification of Herba Taraxaci, Herba Corydalis Bungeanae; HPLC was used for the determination of baicalin. RESULTS: The TLC identification was highly specific and the spots clear and concentrated. The linear range of baicalin was from 0.060～ 0.660?g, r = 0.99991, The recovery of the loading was 97.29%, RSD was 2.13%, respectively. The content of baicalin wasn't less than than 9.0mg per tablet. This method was easy to operate with accurate result, high sensitivity and good reproducibility. CONCLUSION: These methods are able to effectively control the quality of Puqinxiaoyan Tablets.