Antibacterial and osteogenic properties of biomimetic mineralized iodine-loaded coating with micro-nano topography on bone implants / 中华创伤骨科杂志

Objective:To investigate the antibacterial and osteogenic properties of biomimetic mineralized iodine-loaded coating with micro-nano topography on the surface of bone implants.Methods:After the fiber network structure of sodium hydrogen titanate was constructed by alkali thermal reaction on the surface of Ti6Al4V (noted as AT), it was biomimetically mineralized in the modified simulated body fluid to form a micro-nano topology with high specific surface area (noted as AT-CaP), and finally loaded with PVPI to construct a novel antibacterial osseointegration coating (noted as AT-CaP-PVPI). The study was conducted in AT, AT-CaP, and AT-CaP-PVPI groups, in each of which 3 parallel experiments were performed. The morphology and colony counting of Staphylococcus aureus on the coating surface were observed to detect the in vitro antibacterial performance of the coating. Fifteen male SD rats were randomly divided into 3 groups ( n=5): AT, AT-CaP, and AT-CaP-PVPI. After intramedullary injection of Staphylococcus aureus into the lower end of the femur in the SD rats, titanium rods coated with AT, AT-CaP, and AT-CaP-PVPI were inserted into the marrow cavity. The osteogenesis, volume ratio of new bone mass and number of trabeculae on the surface of the femoral implants were compared between the 3 groups 4 weeks after operation. Results:In AT and AT-CaP groups, a large number of bacteria grew in their inherent elliptical or spherical shape on the implant surface and a large number of colonies were seen on the plate; in AT-CaP-PVPI group, the bacteria on the coating surface exhibited membrane deformation and depression, some of them were completely broken and dissolved, and a large number died. There was almost no new bone formation around the implants in AT group; new bone scattered around the implants with discontinuous distribution in AT-CaP group; a great amount of new bone was seen around the implants with even distribution but no signs of infection in AT-CaP-PVPI group. The volume ratio of new bone mass and the number of trabeculae on the implant surface in AT-CaP-PVPI group were 0.453±0.206 and 6.055±0.536, respectively, significantly higher than those in AT group (0.046±0.028 and 1.667±1.249) and AT-CaP group (0.188±0.052 and 3.804±0.889) ( P<0.05). Conclusion:Biomimetic mineralized iodine-loaded coating with micro-nano topography on the surface of bone implants shows good antibacterial and osteogenic properties.

Treatment of humeral shortening complicated with proximal deformity with a monorail external fixator / 中华创伤骨科杂志

Objective:To discuss the key technique and strategy in the treatment of humeral shortening complicated with proximal deformity using a monorail external fixator.Methods:We retro-spectively reviewed 10 patients (8 males and 2 females) with humeral shortening and proximal deformity who had been treated by a monorail external fixator between March 2015 and April 2018.Their mean age was 19.6 years (range, from 15 to 27 years).The humeral shortening was complicated with proximal varus in 8 cases, and with proximal varus plus kyphosis in 2.The affected humeri were 6 to 11 cm (mean, 8.5 cm) shorter than the normal ones.Half-pins were placed onto the lateral upper arm for installation of a unilateral external fixator to correct the proximal humeral deformity immediately after proximal osteotomy.Gradual lengthening was performed after osteotomy in the middle humeral shaft.The limb function was evaluated by the Cattaneo cri-teria.Results:All patients were followed up for 15 to 41 months (mean, 20 months).The length of lengthening ranged from 5 to 12 cm (mean, 7.5 cm).The range of shoulder abduction averaged 160° (from 130° to 180°), much improved than the preoperative average value (90°).Fine ossification was observed in the lengthening part in 9 patients but dysostosis in the lengthening part occurred in one patient who was healed after iliac autografting.No such complications as pin tract infection and radial nerve injury occurred.Ac-cording to the Cattaneo criteria, 9 limbs in 8 cases were excellent and 2 cases good.Conclusions:A monorail external fixator is a reliable choice for treatment of humeral shortening complicated with proximal deformity.To obtain satisfactory clinical effects, it is essential to master the installation technique of external fixator and prevent and control complications as well.

Early transplantation of growth differentiation factor 11-silenced bone marrow stromal cells promotes osteogenesis in necrotic femoral head induced by steroid in rats / 中华创伤骨科杂志

Objective@#To study the effect of growth differentiation factor (GDF) 11-silenced bone marrow stromal cells (BMSCs) on bone regeneration in early steroid-induced osteonecrosis of the femoral head in rats.@*Methods@#After GDF11 expression in BMSCs was inhibited by siRNA, the knockdown efficiency and transfection cytotoxicity were detected. The further experiments both in vitro (n=3) and in vivo (n=8) were divided into 4 groups respectively: blank control group (without any intervention), model group (glucocorticoid treatment), experimental group (siRNA transfection and glucocorticoid treatment) and negative control group (negative control transfection and glucocorticoid treatment). The BMSCs were induced into osteogenic differentiation. Alkaline phosphatase (ALP) staining and alizarin red staining were applied to evaluate the osteogenic differentiation ability of the cells while reverse transcription polymerase chain reaction (qRT-PCR) was employed to detect the relative expression levels of osteogenic markers. The osteogenesis in the necrotic femoral head was evaluated by microCT, H&E staining, immunohistochemistry staining and biomechanical test.@*Results@#No transfection cytotoxicity was found (P>0.05). The ALP staining and alizarin red staining showed that the osteogenic differentiation of BMSCs in the experimental group was better than that in the model group. At the level of mRNA, the relative expression of ALP, runt-related transcription factor (Runx) 2, osteocalcin (OCN) and type Ⅰ collagen (α1) (COL1A1) in the blank control group (1.00±0.09, 1.02±0.23, 1.03±0.30 and 1.02±0.25, respectively) were significantly higher than those in the model group (0.46±0.11, 0.50±0.11, 0.35±0.01 and 0.57±0.02, respectively) but significantly lower than those in the experimental group (1.97±0.30, 0.94±0.19, 1.50±0.18 and 1.28±0.37) (all P<0.05). MicroCT images and quantitative analysis showed that the bone mass in the experimental group was significantly increased compared with the model group (P<0.05). Histological examination showed better bone regeneration and higher expression of Runx2 and COL1 in the necrotic femoral head in the experimental group than in the model group. Improved biomechanical properties were shown in the experimental group compared with the model group (P<0.05).@*Conclusions@#Silence of GDF11 expression may alleviate the inhibitory effect of glucocorticoid on osteogenic differentiation of BMSCs. Early transplantation of GDF11-silenced BMSCs may promote osteogenesis in the necrotic femoral head in rats.

Early transplantation of growth differentiation factor 11-silenced bone marrow stromal cells promotes osteogenesis in necrotic femoral head induced by steroid in rats / 中华创伤骨科杂志

Objective To study the effect of growth differentiation factor (GDF) 11-silenced bone marrow stromal cells (BMSCs) on bone regeneration in early steroid-induced osteonecrosis of the femoral head in rats.Methods After GDF11 expression in BMSCs was inhibited by siRNA,the knockdown efficiency and transfection cytotoxicity were detected.The further experiments both in vitro (n =3) and in vivo (n =8)were divided into 4 groups respectively:blank control group (without any intervention),model group (glucocorticoid treatment),experimental group (siRNA transfection and glucocorticoid treatment) and negative control group (negative control transfection and glucocorticoid treatment).The BMSCs were induced into osteogenic differentiation.Alkaline phosphatase (ALP) staining and alizarin red staining were applied to evaluate the osteogenic differentiation ability of the cells while reverse transcription polymerase chain reaction (qRT-PCR) was employed to detect the relative expression levels of osteogenic markers.The osteogenesis in the necrotic femoral head was evaluated by microCT,H& E staining,immunohistochemistry staining and biomechanical test.Results No transfection cytotoxicity was found (P ＞ 0.05).The ALP staining and alizarin red staining showed that the osteogenic differentiation of BMSCs in the experimental group was better than that in the model group.At the level of mRNA,the relative expression of ALP,runt-related transcription factor (Runx) 2,osteocalcin (OCN) and type Ⅰ collagen (α1) (COL1A1) in the blank control group (1.00 ± 0.09,1.02 ± 0.23,1.03 ± 0.30 and 1.02 ± 0.25,respectively) were significantly higher than those in the model group (0.46±0.11,0.50±0.11,0.35±0.01 and0.57±0.02,respectively) but significantly lower than those in the experimental group (1.97±0.30,0.94±0.19,1.50±0.18 and 1.28 ±0.37) (all P ＜ 0.05).MicroCT images and quantitative analysis showed that the bone mass in the experimental group was significantly increased compared with the model group (P ＜ 0.05).Histological examination showed better bone regeneration and higher expression of Runx2 and COL1 in the necrotic femoral head in the experimental group than in the model group.Improved biomechanical properties were shown in the experimental group compared with the model group (P ＜ 0.05).Conclusions Silence of GDF11 expression may alleviate the inhibitory effect of glucocorticoid on osteogenic differentiation of BMSCs.Early transplantation of GDF11-silenced BMSCs may promote osteogenesis in the necrotic femoral head in rats.

Effects of acupuncture on PI3K/Akt/mTOR signaling pathway in rats with premature ovarian failure / 中国针灸

<p><b>OBJECTIVE</b>To explore the effects of acupuncture and medication on PI3K/Akt/mTOR signaling pathway in rats with premature ovarian failure.</p><p><b>METHODS</b>Ten of fifty SPF-grade female SD rats were randomly selected into a normal group, and the remaining 40 rats were treated with intraperitoneal injection of cyclophospha mide (30 mg/kg) for consecutive 5 days to establish rat model of premature ovarian failure. Thirty five successful rat models were randomly divided into a model group (9 cases), a medication group (9 cases), an acupuncture group A (9 cases) and an acupuncture group B (8 cases). The rats in the model group and normal group did not receive any treatment. The rats in the medication group were treated with intragastric administration of diethylstil bestrol, once a day. The rats in the acupuncture group A and acupuncture group B were respectively treated with acupuncture at different acupoints, twice a day. All the treatment was given for 4 weeks. After the treatment, enzyme-linked immunosorbent assay (ELISA) was applied to test the levels of estradiol (E2), progesterone (P), follicle stimulating hormone (FSH) and luteotropic hormone (LH). The ovarian tissue sample was processed with hematoxylin eosin (HE) staining as well as RNA and protein extraction to test the mRNA expression of estrogen receptor alpha (ERalpha), estrogen receptor beta (ERP), phosphatidylinositol 3-kinase/serine/threonine kinase (PI3K), protein kinase B (Akt) and mammalian target of rapamycin (mTOR).</p><p><b>RESULTS</b>High-dose short-term in- tervention of cyclophosphamide could establish rat model of premature ovarian failure with a successful rate of 87.5%. Compared with the normal group, the vaginal smear in the model group was featured with signs of estro gen deficiency, early-follicle reduction, structural damage to the follicle, and reducing number of mature follicles; the level of E2 was significantly reduced (P<0.05), levels of P, FSH and ILH were increased (all P<0.05), and mRNA expression of estrogen-related ERP3, PI3K, Akt and mTOR were all reduced (all P<0.05). Compared with the model group, the number of mature follicle was increased in the medication group and acupuncture groups, the levels of E2 was obviously increased (all P<0.05). level of FSH was reduced (all P<0.05), and mRNA expression of PI3K, Akt and mTOR all showed an increasing trend (all P<0.05). The differences of each index result between acupuncture groups and medication group were not significant (all P>0.05).</p><p><b>CONCLUSION</b>Acupuncture has certain advantage for the treatment of premature ovarian failure, which achieves similar therapeu tic effect as estrogen; the possible mechanism may be related to up-regulation of gene and protein expression in PI3K/Akt/mTOR signaling pathway.</p>

Analysis of analgesics administration from July 2005 to July 2006 in multicenter emergency rooms in Shenyang area / 中华急诊医学杂志

Objective To analyze analgesics administration from July 2005 to July 2006 in emergency rooms of the First, the Second and Fourth Affiliated Hospitals of China Medical University in order to provide a reference for clinically rational administration. Method The data of 2313 prescriptions with analgesics administered during one-year period were analyzed in many respects including the overview of the prescriptions, the frequency of anal-gesics administration, the system of defined daily doses (DDDs) and drug utilization index (DUI) of narcotic anal-gesics were analyzed, and a survey of 200 patients managed with some of those analgesics was done by using ques-tionaire as they were admitted to and discharged from the emergency room. The pain intensity was evaluated by a 10-point numerical rating scale (NRS). The respondents, excluding the mute or deaf, police custody, victims of domestic violence,mental disorder and age under 14,rated the levels of satisfaction with medication for pain relief. The data of frequency and percentage of the administration of analgeics were analyzed,and the scores of NRS were evaluated with the Paired-samples t -test. Results Most of analgesics were in the form of parenteral route usage, of which anisodamine and bucinnazine were employed in large proportion, and a small number of them was in the form of tablets. Trauma was the commonest cause of pain. Of the narcotic analgesics, meperidine was the most com-monly used analgesics, and its DDDs and DUI were much lower than that of WHO limits. Of the 200 patients, 71.5% patients rated a considerably high satisfaction with scores of (7.47 ±2.21) and (5.00 ± 3. 16) by NRS before and after medication,respectively ( ( -value 23.38,P < 0.01) .The patients presenting pain intensity with a scores of 4 or greater accounted for 57.5% . Conclusions The patients suffering from pain could lie rationally treated in the emergency rooms of those three hospitals. Narcotic analgesics should be cautiously employed, and there is room for improvement in pain management practice in emergency room.

A lamin-like protein gene is down-regulated in human gastric cancer / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To clone human gastric cancer related gene and to analyze its expression profile in gastric mucosal tissues.</p><p><b>METHODS</b>Paired tumor, paratumor and non-tumor specimens from 7 gastric adenocarcinoma patients (male 4, female 3, with average age 51 +/- 18 years) were studied by means of fluorescent differential display reverse transcription polymerase chain reaction (DDRT-PCR). The differentially expressed cDNA bands of interest were cloned and analyzed by Northern blot and in situ hybridization. Thirty cases (male 23 female 7 with average age 59 +/- 8 years) of paired paraffin-embedded gastric tumor and non-tumor tissues were used in in situ hybridization analysis.</p><p><b>RESULTS</b>A gene expressed much lower in 6 out of 7 tested tumor samples than in their normal and paratumor counterparts was identified. It was named GCRG123. Northern blot analysis confirmed the differential expression. Human multiple tissue Northern blot analysis showed that GCRG123 expressed in various adult human tissues including thymus, prostate, testis, ovary, small intestine, colon and peripheral blood leukocyte. Sequence analysis revealed that GCRG123 (GenBank accession number AF454554) was a lamin like protein gene. It had one open reading frame which consisted of 49 amino acids (GenBank accession number AAL61668.1). In situ hybridization analysis showed a high GCRG123 expression level in normal gastric epithelium and pylori glands, but low expression level in tumor as well as dysplasia and most intestinal metaplasia at the paratumor regions.</p><p><b>CONCLUSION</b>A lamin-like protein gene was identified in human gastric mucosa, it is down-regulated in gastric cancer and its precancerous leisions.</p>

Poor prognosis associated with vascular endothelial growth factor(VEGF) and platelet drived endothelial cell growth factor(PD-ECGF) expression in aged patients with gastric carcinoma / 中华老年医学杂志

Objective To investigate the expression of vascular endothelial growth factor (VEGF) and platelet drived endothelial cell growth factor (PD-ECGF) and their influence on the clinicopathological characteristics and the prognosis in aged patients with gastric carcinoma. Methods The expression of VEGF and PD-ECGF were examined with immnunohistochemical technique through preoperative biopsied specimens in 92 cases. Results The expression of VEGF and PD-ECGF were significantly higher in gastric carinoma than in chronic atrophic gastritis. VEGF and PD-ECGF levels in advanced carcinoma were higher compared with early carcinoma( P

A clinicopathological analysis of 22 cases of multiple malignant tumors / 中国医学科学杂志(英文版)

To get a better understanding of the location, pathophysiology, etiology and prognosis of multiple malignant tumors (MPMT), we evaluated the medical records of 22 patients with MPMT. Our results suggested that radiotherapy and chemotherapy might play an important role in the pathogenesis of MPMT and follow-up is important in detecting a secondary primary malignant tumor (PMT) at an early stage. Surgical removal of tumors is the first-choice therapy for MPMT.

Co-expression of immunoglobulin light chain kappa and lambda in gastric carcinoma cell / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To study the expression of immunoglobulin light chain kappa and lambda (Igkappa and Iglambda) in gastric carcinoma cell and their co-expression.</p><p><b>METHODS</b>Igkappa and Iglambda of 22 human gastric carcinoma specimens embedded in paraffin were monitored through immunohistochemical method-LSAB method.</p><p><b>RESULTS</b>Among 22 gastric carcinoma specimens, both Igkappa and Iglambda were positive in 17 (77.3%), only Igkappa was positive in 2 (9.1%), only Iglambda was positive in 1 (4.5%), both Igkappa and Iglambda negative in 2 (9.1%). The expression of Igkappa and Iglambda in human gastric carcinoma cell showed significant close correlation (chi(2) = 5.49, P < 0.05).</p><p><b>CONCLUSION</b>Co-expression of immunoglobulin light chain kappa and lambda in gastric carcinoma cell is common, which suggests that the activation mechanism of immunoglobulin gene in gastric carcinoma cell may be different from that in B-lymphocytes. Study on co-expression of immunoglobulin light chain kappa and lambda in gastric carcinoma is promising.</p>

Direct gastroscopy for detecting gastric cancer in the elderly / 中华医学杂志(英文版)

<p><b>OBJECTIVE</b>To evaluate the safety and effectiveness of direct gastroscopy for detecting gastric cancer.</p><p><b>METHODS</b>Clinical screening by direct gastroscopy was performed for gastric cancer (GC) from September 1985 to July 1998. 3048 elderly people were screened. Their age ranged from 60 to 93 years, and 2034 of the 3084 were followed up.</p><p><b>RESULTS</b>Ninety-two patients with gastric cancer were discovered by gastroscopy, representing 3.02% of the screened population. The rate of early gastric cancer (EGC) was 63.04% (58/92) of all gastric cancers detected. The rate was up to 79.59% (39/49) on follow-up, and was 74.14% (43/51) in asymptomatic patients with gastric cancer. The excision rate was 88.89% for patients with gastric cancer, and 100% for patients with early gastric cancer. The 5-year survival rate was 91.89% for patients with gastric cancer, and 96.30% for patients with early gastric cancer.</p><p><b>CONCLUSION</b>Clinical screening and follow-up by direct gastroscopy in persons over 60 years of age are a safe and effective method for raising the 5-year survival and detection rate of gastric cancer, especially early gastric cancer.</p>

Clinical analysis of recurrence after gastrectomy for early gastric cancer / 解放军医学杂志

Objective To analyze clinicopathological characteristics of recurrence after gastrectomy for early gastric cancer. Methods 308 patients were treated surgically for early gastric cancer from 1983 to 2005. 245 patients were followed up after gastric resection.Clinicopathologic factors were investigated by using univariate methods and multivariate analysis for the possible relationship to recurrence. Results 30 patients developed recurrent disease (median 28 months). The 1-year, 3-year, 5-year, 7-year, 10-year and 15-year recurrent rates were 5.49%, 8.44%, 11.27%, 14.83%, 16.39% and 37.79%, respectively. 13 patients with mucosal gastric cancer developed recurrent disease (median 24 months). The 1-year, 3-year, 5-year, 7-year, 10-year and 15-year recurrent rates were 4.23%, 6.68%, 7.75%, 9.34%, 9.34% and 28.24%, respectively. 17 patients with submucosal gastric cancer developed recurrent disease (median 31 months). The 1-year, 3-year, 5-year, 7-year, 10-year and 15-year recurrent rates were 7.39%, 11.14%, 16.54%, 24.49%, 29.69% and 64.85%, respectively. Cox multivariate analysis showed that submucosal invasion (P=0.044, OR=2.172) was a positive independent risk factor and paracarcinomatous mucosal medium-severe intestinal metaplasia (P=0.047, OR=0.460) was a negative independent risk factor for recurrence. 76.7% (23/30) patients with recurrence did not have indication of resection for a cure, and they did not undergo surgery again. 23.3% (7/30) patients with recurrence had indication for curative resection, and 4 of whom underwent curative resection, but 3 did not because of poor health. Pathological examination after surgery showed that in 3 patients there was early gastric cancer in the remnant stomach without lymph node metastasis, and in one patient there was advanced gastric cancer in remnant stomach with regional lymph node metastasis. The patient with advanced gastric cancer survived for 28 month without detectable tumor. Logistic regression analysis showed paracarcinomatous mucosal medium-severe intestinal metaplasia (P=0.016, OR=17.000) was a positive independent predictor for second radical surgery. The follow-up examinations including endoscopy were performed in 86.7% (26/30) of patients after operation at least every 1-2 years. Conclusion Early gastric cancer patients with submucosal invasion have a high risk of recurrence, and those with paracarcinomatous mucosal medium-severe intestinal metaplasia have a low risk of recurrence. The patients with paracarcinomatous mucosal medium-severe intestinal metaplasia and cancer recurrence are feasible for a curative resection. The follow-up examinations including endoscopy every 1 or 2 years contributed highly to finding an early recurrent cancer in the remnant stomach. But it is not so helpful to increase the possibility of a curative surgery in patients with recurrence and metastasis after gastrectomy.

Multivariate analysis of lymph node metastasis in patients with early gastric cancer / 解放军医学杂志

2cm) and undifferentiated type were significantly related to lymph node metastasis in submucosal cancer. No lymph node metastasis was observed in 25 patients with submucosal invasion who showed none of the three risk factors, whereas 72.7% (8/11) of patients with all the three factors had lymph node metastasis. Conclusion The tumor size and lymphatic vessel involvement are related with lymph node metastasis in mucosal cancer. Poor differentiation, tumor size and lymphatic vessel involvement are related with lymph node metastasis in submucosal cancer.

Preparation of gastric adenocarcinoma related cDNA microarray / 解放军医学杂志

Objectives To prepare the gastric adenocarcinoma related cDNA microarray and to screen differentially expressed genes between human gastric adenocarcinoma and normal gastric mucosa. Methods A set of cDNA microarray slides containing a duplicate set of 576 cDNA spots. These selected clones have been verified in sequence. The PCR products of human genes were spotted onto a chemical-material-coated glass plates in array. DNAs were fixed on the glass plate after series of treatment. Both RNAs from the gastric adenocarcinoma and normal gastric mucosa were reversely transcribed to the cDNAs with the incorporation of fluorescent dUTP to prepare the hybridization probes. The mixed probes were hybridized to the cDNA microarray. After high-stringent washing, the cDNA microarray was scanned for fluorescent signals to display differences between two tissues. Results Gastric adenocarcinoma related cDNA microarray slides contained 111 differentially expressed genes of gastric adenocarcinoma, 37 known genes related to gastric carcinogenesis (such as p21ras, p53, etc.) and control genes. The results of 10 couples of hybridizations to the gastric adenocarcinoma related cDNA microarray slides with Cy3-labeled cDNA probe from normal gastric tissue and Cy5-labeled cDNA probe from gastric adenocarcinoma showed analyzed spots uniformity background and clear signal. Among the 576 target genes, the expression level of 48 genes differed between gastric adenocarcinoma and normal mucosa in 5 or more samples (eight genes differed in all samples). There were 39 down-regulated and 9 up-regulated genes in gastric adenocarcinoma. Conclusion Gastric adenocarcinoma related cDNA microarray slides were successfully developed. The results of the experiments showed that gastric adenocarcinoma related cDNA microarray slides are reliable, sensitive and reproducible.

Tissue microarray for detecting the expression of PCNA,Cyclin D1 and P27 during the course of gastric carcinogenesis / 解放军医学杂志

Objective To detect the expressions of PCNA, Cyclin D1 and P27, which were related to cell proliferation and cell cycle in gastric carcinoma and precancerous lesion of gastric mucosa. Methods Tissue microarray technique was applied to construct a tissue microarray containing gastric carcinoma, precancerous lesions and normal gastric mucosa. Diagnosis of the tissue in the microarray was pathologically confirmed. Immunohistochemical method was used to assess the expression of PCNA, Cyclin D1 and P27 in all the samples of tissue microarray. Results The expressions of PCNA and Cyclin D1 in gastric carcinoma, gastric hyperplasia, and intestinal metaplasia tissues were higher than that in normal gastric mucosa. The expressions of P27 in gastric hyperplasia, intestinal metaplasia and gastric carcinoma tissues were lower than that in normal gastric mucosa. Conclusion Gastric carcinoma, precancerous lesion and normal gastric mucosa tissues present different degrees of expression of PCNA, Cyclin D1 and P27, either with a increasing trend or a decreasing trend. These changes in expression are all closely related to gastric carcinogenesis.

Studies on the function of FBG2 in gastric cancer cell lines / 解放军医学杂志

Objective To investigate the influence of FBG2 on the growth, proliferation, apoptosis, infiltration and cell cycle of the gastric cancer line MKN45. Methods A critical component ubiquitin-protein ligase complex, FBG2 cDNA, was subcloned into a constitutive vector pcDNA3.1, followed by transfection in MKN45 by using liposome. Then stable expression clones were selected and appraised. The apoptosis and cell cycles were detected using flow cytometry. The growth and proliferation were analyzed by plotting cell growth curves and colony formation assay respectively. The ability of infiltration was tested using cancer cell migration assay. The MKN-FBG2 group and two control groups were included in the study. Results MKN-FBG2 grew faster than MKN45 and MKN-PC. The cell counts of MKN-FBG2 on the fourth, fifth, sixth and seventh day were significantly larger than that of others (P

Construction of gastric cancer related gene GCRG213 eukaryotic expression vector and its effect on growth of gastric cancer cells / 解放军医学杂志

Objective To investigate the effect of gene GCRG213 transfection (sense, anti-sense) on growth of gastric cancer cell MKN45. Methods The sense and anti-sense fragment of GCRG213 were obtained by PCR. They were cloned into eukaryotic expression vector pcDNA3.1(+). The recombinant plasmid pcDNA3.1-a, pcDNA3.1-b and the vector pcDNA3.1 were transfected separately into MKN45 cells conducted by lipofectamine~ TM 2000. Expression of GCRG213 was assessed with semi-quantitative RT-PCR and Western Blot. The growth graph was plotted by the methods of cell counting of three steady transfected cells. FACS was used to determine the cell cycle, and Annexin V FITC/PI bi-labeling method was used to determine the cell apoptosis. Results By sequencing, sense GCRG213 and anti-sense GCRG213 were proved to be successfully cloned into pcDNA3.1. Transfecting the sense vector (pcDNA3.1-a) into the MKN45 significantly increased the expression of GCRG213, both in mRNA level and protein level. Transfecting the anti-sense vector (pcDNA3.1-b) into the MKN45 significantly decreased the expression of GCRG213, both in mRNA level and protein level. The growth of pcDNA3.1-a transfected cells was faster than that of vector transfected cells, and the cell apoptosis decreased. But the growth of pcDNA3.1-b transfected cells was slower in multiplication than that of vector transfected cells, and the cell apoptosis was increased. Conclusion Stable transfection showed that GCRG213 promoted cell to grow, inhibited the cell apoptosis. GCRG213 might be a new promoter to tumor.

Expression of gastric cancer related gene GCRG213 in E.coli and purification of recombinant protein / 解放军医学杂志

Objective To express gastric cancer related gene GCRG213 by using thioredoxin fusion expression system, and to prepare human GCRG213 fusion protein. Methods GCRG213 cDNA with complete open reading frame was amplified by PCR from plasmid pGEM-T, and then was cloned into thioredoxin fusion expression vector pET102/D-TOPO. The recombinant plasmid was further transformed into E.coli BL21 strain. After induction with IPTG, the thioredoxin/GCRG213 fusion protein was expressed in E.coli. The product was obtained by means of direct purification from a denaturing polyacrylamide gel. Results SDS-PAGE analysis showed the thioredoxin/GCRG213 fusion protein with relative molecule mass of 29.4kDa was highly expressed. The thin layer gel scanning analysis showed that the yield of GCRG213 fusion protein was 28.7% of the total bacterial protein. The product was obtained with a purity of about 100% by means of direct purification from a denaturing polyacrylamide gel. Conclusion The thioredoxin/GCRG213 fusion protein was successfully expressed in E.coli and the product with high purity was obtained, which laid the foundation for the function research and antibody preparation hereafter.

Preparation and identification of polyclonal antibody against gastric cancer-related protein GCRG224 / 解放军医学杂志

Objective To prepare a polyclonal antibody against gastric cancer-related protein GCRG224.Methods The thioredoxin/GCRG224 fusion protein was expressed in E.coli.A polyclonal antibody against GCRG224 was obtained by immunizing a rabbit with the purified GCRG224 protein.The titer and specificity of the antibody were determined by ELISA and Western-blot,respectively.Results The thioredoxin/GCRG224 fusion protein with relative molecular mass of 16.8kD was over-expressed in E.coli.The purity of expressed products directly purified from a denaturing polyacrylamide gel was about 100%.The polyclonal antibody against GCRG224 was obtained.The ELISA titer of antiserum against GCRG224 was about 1∶256 000.Western blot analysis showed that the antiserum could bind to the expressed fusion protein specifically.Conclusion The polyclonal antibody against GCRG224 has been successfully prepared,which lays the foundation for further study on the biological function and the possible role of the GCRG224 in the development of gastric carcinoma.