Family-based Helicobacter pylori infection control and management strategy and screen-and-treat strategy are highly cost-effective in preventing multiple upper gastrointestinal diseases in Chinese population at national level.

BACKGROUND: The overall benefits of the newly introduced family-based Helicobacter pylori (H. pylori) infection control and management (FBCM) and screen-and-treat strategies in preventing multiple upper gastrointestinal diseases at national level in China have not been explored. We investigate the cost-effectiveness of these strategies in the whole Chinese population. MATERIALS AND METHODS: Decision trees and Markov models of H. pylori infection-related non-ulcer dyspepsia (NUD), peptic ulcer disease (PUD), and gastric cancer (GC) were developed to simulate the cost-effectiveness of these strategies in the whole 494 million households in China. The main outcomes include cost-effectiveness, life years (LY), quality-adjusted life year (QALY), and incremental cost-effectiveness ratio (ICER). RESULTS: When compared with no-screen strategy, both FBCM and screen-and-treat strategies reduced the number of new cases of NUD, PUD, PUD-related deaths, and the prevalence of GC, and cancer-related deaths. The costs saved by these two strategies were $1467 million and $879 million, quality-adjusted life years gained were 227 million and 267 million, and life years gained were 59 million and 69 million, respectively. Cost-effectiveness analysis showed that FBCM strategy costs -$6.46/QALY and -$24.75/LY, and screen-and-treat strategy costs -$3.3/QALY and -$12.71/LY when compared with no-screen strategy. Compared to the FBCM strategy, the screen-and-treat strategy reduced the incidence of H. pylori-related diseases, added 40 million QALYs, and saved 10 million LYs, but at the increased cost of $588 million. Cost-effectiveness analysis showed that screen-and-treat strategy costs $14.88/QALY and $59.5/LY when compared with FBCM strategy. The robustness of the results was also verified. CONCLUSIONS: Both FBCM and screen-and-treat strategies are highly cost-effective in preventing NUD, PUD, and GC than the no-screen strategy in Chinese families at national level. As FBCM strategy is more practical and efficient, it is expected to play a more important role in preventing familial H. pylori infection and also serves as an excellent reference for other highly infected societies.

One-pot catalyst-free synthesis of benzopyrroxazine derivatives by a mutually activated sequential annulation process.

A one-pot catalyst-free reaction of o-hydroxyaryl azomethine ylides, vinyl pyridines and paraformaldehyde for the synthesis of benzopyrroxazines is reported, which offers a straightforward and atom-economical procedure for the preparation of benzopyrroxazine derivatives in moderate to excellent yields under mild conditions. A self-catalyzed [3 + 2] annulation through a mutual activation method and a sequential non-catalyzed [5 + 1] annulation process contribute to this strategy. The corresponding control experiments have been conducted to reveal the mechanism of this reaction.

Heterogeneous immunogenomic features and distinct escape mechanisms in multifocal hepatocellular carcinoma.

BACKGROUND & AIMS: The presence of multifocal tumors, developed either from intrahepatic metastasis (IM) or multicentric occurrence (MO), is a distinct feature of hepatocellular carcinoma (HCC). Immunogenomic characterization of multifocal HCC is important for understanding immune escape in different lesions and developing immunotherapy. METHODS: We combined whole-exome/transcriptome sequencing, multiplex immunostaining, immunopeptidomes, T cell receptor (TCR) sequencing and bioinformatic analyses of 47 tumors from 15 patients with HCC and multifocal lesions. RESULTS: IM and MO demonstrated distinct clonal architecture, mutational spectrum and genetic susceptibility. The immune microenvironment also displayed spatiotemporal heterogeneity, such as less T cell and more M2 macrophage infiltration in IM and higher expression of inhibitory immune checkpoints in MO. Similar to mutational profiles, shared neoantigens and TCR repertoires among tumors from the same patients were abundant in IM but scarce in MO. Combining neoantigen prediction and immunopeptidomes identified T cell-specific neoepitopes and achieved a high verification rate in vitro. Immunoediting mainly occurred in MO but not IM, due to the relatively low immune infiltration. Loss of heterozygosity of human leukocyte antigen (HLA) alleles, identified in 17% of multifocal HCC, hampered the ability of major histocompatibility complex to present neoantigens, especially in IM. An integrated analysis of Immunoscore, immunoediting, TCR clonality and HLA loss of heterozygosity in each tumor could stratify patients into 2 groups based on whether they have a high or low risk of recurrence (p = 0.038). CONCLUSION: Our study comprehensively characterized the genetic structure, neoepitope landscape, T cell profile and immunoediting status that collectively shape tumor evolution and could be used to optimize personalized immunotherapies for multifocal HCC. LAY SUMMARY: Immunogenomic features of multifocal hepatocellular carcinoma (HCC) are important for understanding immune-escape mechanisms and developing more effective immunotherapy. Herein, comprehensive immunogenomic characterization showed that diverse genomic structures within multifocal HCC would leave footprints on the immune landscape. Only a few tumors were under the control of immunosurveillance, while others evaded the immune system through multiple mechanisms that led to poor prognosis. Our study revealed heterogeneous immunogenomic landscapes and immune-constrained tumor evolution, the understanding of which could be used to optimize personalized immunotherapies for multifocal HCC.

Topical Review: Causes of Refractive Error After Silicone-oil Removal Combined with Cataract Surgery.

SIGNIFICANCE: This review summarizes the main factors of refractive error after silicone oil removal combined with cataract surgery.The post-operative refractive results of silicone oil removal combined with cataract surgery are closely related to the patient's future vision quality. This report summarizes the factors that influence the difference between the actual post-operative refractive power and the pre-operatively predicted refractive power after silicone oil removal combined with cataract surgery, including axial length, anterior chamber depth, silicone oil, commonly used tools for measuring intraocular lens power, and intraocular lens power calculation formulas, among others. The aim of the report is to assist clinical and scientific research on the elimination of refractive error after silicone oil removal combined with cataract surgery.

Circumventing intratumoral heterogeneity to identify potential therapeutic targets in hepatocellular carcinoma.

BACKGROUND & AIMS: Identifying target genetic mutations in hepatocellular carcinoma (HCC) for therapy is made challenging by intratumoral heterogeneity. Circulating cell-free DNAs (cfDNA) may contain a more complete mutational spectrum compared to a single tumor sample. This study aimed to identify the most efficient strategy to identify all the mutations within heterogeneous HCCs. METHODS: Whole exome sequencing (WES) and targeted deep sequencing (TDS) were carried out in 32 multi-regional tumor samples from five patients. Matched preoperative cfDNAs were sequenced accordingly. Intratumoral heterogeneity was measured using the average percentage of non-ubiquitous mutations (present in parts of tumor regions). Profiling efficiencies of single tumor specimen and cfDNA were compared. The strategy with the highest performance was used to screen for actionable mutations. RESULTS: Variable levels of heterogeneity with branched and parallel evolution patterns were observed. The heterogeneity decreased at higher sequencing depth of TDS compared to measurements by WES (28.1% vs. 34.9%, p<0.01) but remained unchanged when additional samples were analyzed. TDS of single tumor specimen identified an average of 70% of the total mutations from multi-regional tissues. Although genome profiling efficiency of cfDNA increased with sequencing depth, an average of 47.2% total mutations were identified using TDS, suggesting that tissue samples outperformed it. TDS of single tumor specimen in 66 patients and cfDNAs in four unresectable HCCs showed that 38.6% (26/66 and 1/4) of patients carried mutations that were potential therapeutic targets. CONCLUSIONS: TDS of single tumor specimen could identify actionable mutations targets for therapy in HCC. cfDNA may serve as secondary alternative in profiling HCC genome. LAY SUMMARY: Targeted deep sequencing of single tumor specimen is a more efficient method to identify mutations in hepatocellular carcinoma made from mixed subtypes compared to circulating cell-free DNA in blood. cfDNA may serve as secondary alternative in profiling HCC genome. Identifying mutations may help clinicians choose targeted therapy for better individual treatments.

Studies on anti-hepatoma activity of Annona squamosa L. pericarp extract.

This study investigated the anti-hepatoma activity of different extracts from A. squamosa pericarps, phytochemistry of the ethyl acetate (EtOAc) fraction and possible anti-hepatoma mechanism of active constituents. The anti-hepatoma activity of different extracts from A. squamosa pericarps were evaluated by MTT assay against SMMC-7721 cells in vitro and verified by using H22 xenografts bearing mice. Phytochemical investigation of the active pericarp extract was carried out. The pro-apoptosis and cycle arrest effects of active constituents were observed by fluorescent microscope and flow cytometry. Western blot assay was conducted to find the possible anti-hepatoma mechanisms of active constituents. The result showed that EtOAc extract was the active fraction. Two ent-kaurane diterpenoids, named ent-kauran-16-en-19-oic acid and ent-kauran-15-en-19-oic acid, were isolated from the active EtOAc fraction. The pro-apoptosis and G1 phase arrest effects of these diterpenoids were found. Western blot assay showed that ent-kauran-16-en-19-oic acid could activate caspase-3,-8,-9, up-regulate of Bax and down-regulate of Bcl-2.

Application of a sensitive and accurate LC-MS/MS method for determination of dryocrassin ABBA in rat plasma for a bioavailability study.

A sensitive and accurate liquid chromatography-tandem mass spectrometry method was developed and validated for the determination of dryocrassin ABBA, a potential active component isolated from Dryopteris crassirhizoma, in rat plasma. Chromatographic separation was achieved on a Zorbax SB-C18 column (50 × 2.1 mm, 1.8 µm), with elution consisting of eluent (A) 10 mm ammonium acetate in methanol containing 0.1% formic acid and (B) 10 mm ammonium acetate in water containing 0.1% formic acid (A:B = 99:1, v/v) at a flow rate of 0.3 mL/min. Multiple reaction monitoring mode was used to monitor the precursor-product ion transitions of m/z 819.3 → 403.4 for dryocrassin ABBA and m/z 426.2 → 409.2 for internal standard. This assay exhibited a good linearity with a correlation coefficient >0.99 and showed no endogenous interference with the analyte and internal standard. The lower limit of quantification of dryocrassin ABBA was 4 ng/mL in 50 µL of rat plasma. The method was successfully applied in the pharmacokinetic study of dryocrassin ABBA in rats after intravenous (2.35 mg/kg) and oral (23.5 mg/kg) doses of dryocrassin ABBA. The oral bioavailability (F) of dryocrassin ABBA was estimated to be 50.1%. Our study is the first to clarify the pharmacokinetic behaviors of dryocrassin ABBA in animals.

Treatment of a non-typical hepatic pseudolesion complicated by greatly elevated alpha fetoprotein: case report and literature review.

BACKGROUND: Hepatic pseudolesions detected by helical computed tomography (CT) are not rare, but it is difficult to make a final diagnosis when the hepatic lesion is complicated by the presence of greatly elevated alpha fetoprotein (AFP). Clinical treatment of non-typical hepatic pseudolesions complicated by greatly elevated AFP should confirm the diagnosis and minimize trauma. CASE PRESENTATION: Non-invasive procedures including ultrasonography, CT, and micro-invasive digital subtraction angiography could not safely differentiate this lesion from a malignant focus when it was complicated by greatly elevated AFP. Laparoscopic hepatectomy was performed, and pathological analysis showed chronic hepatitis, nodular regenerative hyperplasia, focal nodular hyperplasia of the liver, and mild vascular malformation. The tissue was HbsAg(-), HbcAg(-), and AFP(+). CONCLUSION: Heightened awareness of hepatic pseudolesion complicated by primarily elevated AFP will help physicians avoid unnecessary invasive procedures. Hepatic biopsy is inevitable because of greatly elevated AFP. For suspected hepatic pseudolesion with elevated AFP, needle-core biopsy and follow-up surveillance instead of hepatectomy are recommended to find the source of AFP and make a final diagnosis of pseudolesion.

TNF-α, HGF and macrophage in peritumoural liver tissue relate to major risk factors of HCC Recurrence.

BACKGROUND/AIMS: Hepatocyte growth factor (HGF) responds to tumor necrosis factor- α (TNF- α) secreted by macrophages and has been suggested to function in malignant metastasis. This study was performed to shed a light on the complicated relation between intrahepatic macrophages, TNF- α, HGF and hepatocellular carcinoma (HCC) microvascular invasion (MVI) and pathological grade. METHODOLOGY: Forty-eight HCC cases were divided into three groups according to pathological grade: poorly-differentiated (PD), moderately-differentiated (MD) and well-differentiated (WD) groups. Each group was divided into subgroups based on the condition of MVI. Macrophages were counted, and TNF- α and HGF were tested in all specimens by immunohistochemical analysis and RT-PCR. RESULTS: In peritumoural liver tissue, MVI had more HGF (t=8.22, p<0.05) and less TNF- α (t=4.20, p<0.05) than non-MVI. With MVI, PD and MD had more TNF- α (t=3.30, p<0.05and t=2.82, p<0.05) than WD in peritumoural tissue. CONCLUSIONS: Overexpressed TNF- α and HGF in microenvironment were related to poor differentiation and microvascular invasion of HCC. We suggest that inflammation of hepatic microenvironment promote pathological degradation and microvascular invasion of HCC.

[Study on propofol and fentanyl administrated via common carotid artery in rabbits].

OBJECTIVE: To compare the pharmacodynamic differences of common carotid artery administration with ear vein administration of propofol and fentanyl in rabbits. METHODS: Sixty New Zealand white rabbits were randomly divided into four groups(n = 15):PvFv, PvFa, PaFv and PaFa groups. Propofol 30 mg×kg(-1)×h(-1) and fentanyl 2 µg×kg(-1)×h(-1)were administrated via the ear vein or the common carotid artery. The outcomes were recorded, including the time of consciousness loss and recovery, to electrocerebral silence, dose of propofol and fentanyl, mean arterial pressure, heart rate, respiration rate and SpO2. RESULTS: (1) None of rabbits appeared breathing to be depressed seriously in group PaFa, while respiratory in the other groups were significantly depressed. (2) The dosage of propofol and fentanyl of group PaFa was significantly less than the other groups (P < 0.05). (3) The time of consciousness loss and recovery of group PaFa were shorter than the other three groups. CONCLUSION: Compared to drugs infusion via the ear vein, infusion of propol and fentanyl via the common carotid artery is more advantageous in some aspects, such as rapid anesthesia induction and recovery, smaller dose, and smaller impact on the hemodynamic and respiratory.

Chemical composition and anti-hepatoma effect of Annona squamosa L. pericarp oil.

In this study, an optimal method used to extract Annona squamosa pericarp oil (ASPO) was established according to the response surface model. The yield of ASPO was 1.45%. 8 fatty acids were identified from ASPO by GC-MS. Among them, (9Z)-9-Octadecenoic acid was abundant and accounted for 49.65%. The anti-hepatoma activities of ASPO were investigated against SMMC-7721 cell line in vitro and H22 cell line in vivo. Proteins associated with apoptosis in tumour tissue were quantified by western blot assay. The result revealed that ASPO had significant anti-hepatoma activities with IC50 value of 15.96 µg/mL in vitro and tumour inhibition rate of 54.14% at 50 mg/kg dose in vivo. Protein analysis showed that ASPO activated apoptosis by down-regulating Bcl-2, up-regulating Bax, cleaving caspase 9, cleaving caspase 8 and cleaving caspase 3 proteins. The possible mechanisms of apoptosis induced by ASPO were related to Fas/FasL/caspase-8/caspase-3 and Bcl-2/bax/caspase-9/caspase-3 pathways.

[The diagnostic value of digital subtraction angiography in detecting small intestinal bleeding].

OBJECTIVE: To determine the diagnostic value of digital subtraction angiography (DSA) in detecting small intestinal bleeding. METHODS: DSA was performed in 39 patients with suspected small intestinal bleeding. The sensitivity, specificity and accuracy of DSA were evaluated in the 25 patients who accepted surgical treatment. RESULTS: Among the 39 patients, 33 came out with positive in DSA. Eight patients were given interventional embolization successfully. The sensitivity, specificity and accuracy of DSA in detecting small intestinal bleeding were 91.7%, 100% and 92% respectively. The pathological results confirmed 77.3% of etiological diagnosis by DSA. CONCLUSION: DSA is an effective method for detecting small intestinal hemorrhage. The accurate diagnosis can guide the intervention with embolization.

Treatment of hepatic venous outflow stenosis after living donor liver transplantation by insertion of an expandable metallic stent.

BACKGROUND: Hepatic venous outflow stenosis is an uncommon but serious complication after right lobe living donor liver transplantation (LDLT). Failure to recognize and treat this complication early can result in graft failure and even death. The early diagnosis and management of hepatic venous outflow stenosis has become an important issue. METHOD: We report a case with this complication treated by endovascular stent placement in the early period after right lobe LDLT and review related reports to explore the possible mechanism. RESULTS: A 44-year-old man with end-stage hepatitis B liver cirrhosis underwent right lobe LDLT. On postoperative day 13, his liver function deteriorated and he developed refractory ascites for maximal diuretic therapy. Hepatic venography showed a stenosis with an element of torsion of the venous drainage proximal to the anastomosis of the right hepatic venous orifice and inferior vena cava. The stenosis was successfully treated by insertion of an expandable metallic stent. CONCLUSIONS: The result demonstrates that stent placement for stenosis is safe and effective. We suspect that rapid hypertrophy of an asymmetric right lobe graft may result in hepatic venous outflow stenosis caused by twisting or external compression of the hepatic veins.

Asymptomatic left posterior mediastinal functional paraganglioma: A case report.

RATIONALE: Paraganglioma refers to a set of neuroendocrine tumors derived from the chromaffin cells of the adrenal diplomatic ganglion. Paragangliomas can be classified as functional or nonfunctional based on the ability to synthesize and release catecholamines. PATIENT CONCERNS: We report a 47-year-old man with a functional paraganglioma in the left posterior mediastinum and highlight the key elements of management of mediastinal paragangliomas. DIAGNOSES: A left posterior mediastinal mass was found by computed tomography (CT) scan and Chest-enhanced CT. Preoperative ultrasound-guided biopsy suggested the possibility of a paraganglioma. A diagnosis of paraganglioma was established by immunohistochemistry. INTERVENTIONS: The patient underwent single-stage resection of the lesion via left thoracotomy after preoperative oral α-adrenoceptor (phenoxybenzamine) therapy and intravenous fluid resuscitation for two weeks. OUTCOMES: The postoperative period was uneventful. The patient exhibited no abnormal blood pressure or recurrence during the 12-month follow-up period. LESSONS SUBSECTIONS AS PER STYLE: Pathological examination alone cannot determine whether it was a benign or malignant paraganglioma, which can be determined by pathological examination combined with distant metastasis. Long-term follow-up is required to assess the treatment effect.

Maize productivity and soil properties in the Loess Plateau in response to ridge-furrow cultivation with polyethylene and straw mulch.

Ridge-furrow with full film mulching (RFFM) is widely used in the Loess Plateau (LP) to increase maize yield. However, continuous RFFM application may cause excessive depletion of soil organic carbon (SOC) and soil water storage (SWS). The present study tested four production systems, namely, (1) RFFM; (2) ridge-furrow with polyethylene film and straw mulching (RFFSM); (3) non-contoured seedbed with film mulching (FFM); and (4) non-contoured seedbed without mulching (CK) in 2013 and 2014 to identify an optimal technique to increase maize yield yet minimizing the negative effects. SWS under RFFSM was significantly higher by 5.4% and 13.4% compared to RFFM and CK, respectively. The changes in SOC were -0.2, -0.2, and -0.4 g·kg-1 for RFFM, FFM, and CK, respectively, and 0.3 g·kg-1 for RFFSM. Increased root residue and extra external carbon input to soil under RFFSM directly contributed to SOC recovery. RFFSM had a comparable grain yield but higher water use efficiency compared to RFFM. The combination of RFFSM is promising for improving SOC stocks, water storage, and maize productivity.

The inhibitory effects of cisplatin-radiation combination treatment on malignant osteosarcoma MG-63 cells and BRCA1-p53 pathways are more efficient than single treatments.

The poor prognosis of patients with osteosarcoma remains a persistent problem, in particular for patients with unresectable tumors or metastasis. Therefore, combination of radiotherapy and chemotherapy has been considered for patients with metastasis or recurrence, patients unsuitable for surgery and patients refusing surgery. The present study aimed to investigate the effect of the combined treatment with cisplatin and radiation therapy on the biological characteristics of the osteosarcoma cell line MG-63 and the breast cancer 1 (BRCA1)-associated signaling pathways. Cell proliferation was determined using Cell Counting kit-8 assay, and cell apoptosis and cell cycle were assessed by flow cytometry. Cell migration was examined by Transwell assay. The mRNA and protein expression levels of candidate genes, including BRCA1 and p53, were determined by reverse transcription-quantitative PCR and western blotting, respectively. The results demonstrated that combined treatment with radiation and cisplatin significantly inhibited MG-63 cell proliferation compared with radiation or cisplatin treatment alone. Furthermore, radiation, cisplatin or the combined treatment with radiation and cisplatin increased the apoptosis rate of MG-63 cells, which resulted in G2 phase arrest, and significantly decreased the migratory capacity of MG-63 cells. In addition, the apoptosis rate of MG-63 cells following combined radiation and cisplatin treatment was higher compared with the cisplatin group, but lower compared with the radiation group. Furthermore, combined treatment with radiation and cisplatin decreased the mRNA and protein expression levels of BRCA1 and p53. Additionally, combined treatment with radiation and cisplatin had a more potent inhibitory effect on p53 expression than on BRCA1 expression. In addition, combination of radiation and cisplatin had a higher inhibitory effect on Bax protein level and a higher inductive effect on Bcl-2 protein level compared with treatments with radiation and cisplatin alone. The results demonstrated that combined treatment of radiation and cisplatin exhibited superior therapeutic effects on osteosarcoma MG-63 cells compared with radiation or cisplatin treatment alone, which may be mediated by the BRCA1-p53 signaling pathway.

[Influence of recipient pretransplant MELD-AS score and graft size on the outcome of adult-to-adult living donor liver transplantation].

OBJECTIVE: To investigate the relationship between the pretransplant condition of recipient, graft size and the outcomes of recipient after adult-to-adult living donor liver transplantation. METHODS: From January 2002 to January 2007, 70 patients underwent adult-to-adult living donor liver transplantation (ALDLT) in West China Hospital. There were 67 patients received right lobe graft without middle hepatic vein (MHV) and 3 patients received dual grafts. The patients were followed up until January 2008. The recipients were stratified according to pretransplant MELD-AS score and graft size. The outcomes of these patients were analyzed retrospectively. RESULTS: Recipient 1-year survival rate and complication rate were 87.1% and 34.3%, respectively. For the patients with MELD-AS score from 1 to 13, the survival rates of large graft and small graft were 100.0% and 83.3%, the complication rates were 14.3% and 16.7%; for those MELD-AS score between 14-24, the survival rates of large graft and small graft were 94.4% and 75.0%, the complication rates were 38.9% and 50.0%; for those MELD-AS scores were 25 or higher, the survival rates of large graft and small graft were 77.8% and 25.0%, the complication rates were 52.9% and 50.0%. Statistical significant difference of survival between large graft group and small graft group was found among the recipients with MELD-AS score > or =25. CONCLUSION: The recipient survival is influenced simutanously by pretransplant MELD-AS score of recipient and graft size influence. Further more, large graft improve the survival in recipients whose MELD-AS score were 25 or higher.

[The possible causes of serious central nervous system complications occurring after liver transplantation].

OBJECTIVE: To study the classification, possible causes, cure and prevention measures of serious central nervous system (CNS) complications occurring early following liver transplantation. METHODS: The clinical material records of 347 patients undergoing the orthotopic liver transplantation from July 2001 to July 2006 were analyzed retrospectively. The perioperation risk factors of CNS complications, which might be the primary liver disease, serum sodium level, magnesium level, fluctuation degree of plasma osmolality and serum sodium, function of blood clotting, etc. , were analyzed between patients with serious CNS complications and without neurological complications. RESULTS: A total of 71 (20. 46%) neurological symptoms appeared in 347 postoperative patients who included 6 cases (1. 73%) with serious CNS complications such as cerebral edema, cerebral hemorrhage and central pontine myelinolysis, and of 6 cases, 5 patients died. The morbidity of early serious complications of CNS was higher in patients with decompensate cirrhosis and/or hypersplenism. Compared with the group of no neurological complications, two patients with cerebral hemorrhage suffered from preoperative hypersplenism and worse dysfunction of blood clotting (P<0. 05); patients with developed CPM and cerebral oedema had serious hyponatremia history before operation, and intensely fluctuated plasma osmolality showed before and after operation. CONCLUSION: The onset of serious complications of CNS in recipients of liver transplantation may be intimately related to the primary liver disease, and be associated with chronic hyponatremia, rapid correction of serum sodium concentration, intense increase of plasma osmolality and no prompt rectification of the function of blood clotting during perioperation.

[Isolation and purification of BMScs of GFP transgenic mouse using the method of adhering to cuture plastic in different time].

OBJECTIVE: To adopt the method of adhering to culture plastic in different time for cultivating and purifying BMSCs of green fluorescent protein (GFP) transgenic mice. METHODS: Bone marrow cells isolated from GFP transgenic mice are directly planted in culture flask and an exchange of the total volume medium is made at different time. Then the cells adhering to culture plastic are differently counted according to the cell types and are examined by immunohistochemistry using the antibodies of CD44, CD45 and CD54 in three days. Moreover, the cells after the exchange of the total volume medium in 4 hours, 8 hours and 24 hours are selected and successively subcultured down to the fifth passage. Then the result of amplification is calculated and the cells are examined by immunohistochemistry using the antibodies of CD44, CD45 and CD54. RESULTS: With the extending of the time for the first exchange of medium, the density of cells adhering to culture plastic increased accordingly, but the BMSCs proportion decreased. The cells after first exchange of medium in 4 hours had high BMSCs proportion but low BMSCs density, and the cells in 24 hours had high BMSCs density and low BMSCs proportion. However, the cells in 8-10 hours had high BMSCs density and also high BMSCs proportion. The subcultured BMSCs could stably express GFP. CONCLUSION: The method of adhering to culture plastic in different time for cultivating and purifying BMSCs of GFP transgenic mice is effective. It is suitable to make the first exchange of total volume medium in 8-10 hours. The subcultured cell has the capacity for amplification and will probably be a seed cell for the research of tissue engineering and gene therapy.