Mechanistic insights into AMPK-SIRT3 positive feedback loop-mediated chondrocyte mitochondrial quality control in osteoarthritis pathogenesis.

Osteoarthritis (OA) is a major cause of disability in the elderly population and represents a significant public health problem and socioeconomic burden worldwide. However, no disease-modifying therapeutics are currently available for OA due to an insufficient understanding of the pathogenesis of this disability. As a unique cell type in cartilage, chondrocytes are essential for cartilage homeostasis and play a critical role in OA pathogenesis. Mitochondria are important metabolic centers in chondrocytes and contribute to cell survival, and mitochondrial quality control (MQC) is an emerging mechanism for maintaining cell homeostasis. An increasing number of recent studies have demonstrated that dysregulation of the key processes of chondrocyte MQC, which involve mitochondrial redox, biogenesis, dynamics, and mitophagy, is associated with OA pathogenesis and can be regulated by the chondroprotective molecules 5' adenosine monophosphate-activated protein kinase (AMPK) and sirtuin 3 (SIRT3). Moreover, AMPK and SIRT3 regulate each other, and their expression and activity are always consistent in chondrocytes, which suggests the existence of an AMPK-SIRT3 positive feedback loop (PFL). Although the precise mechanisms are not fully elucidated and need further validation, the current literature indicates that this AMPK-SIRT3 PFL regulates OA development and progression, at least partially by mediating chondrocyte MQC. Therefore, understanding the mechanisms of AMPK-SIRT3 PFL-mediated chondrocyte MQC in OA pathogenesis might yield new ideas and potential targets for subsequent research on the OA pathomechanism and therapeutics.

Correlations between inflammatory cytokines, muscle damage markers and acute postoperative pain following primary total knee arthroplasty.

BACKGROUND: Despite the success of total knee arthroplasty (TKA) in reducing knee pain and improving functional disability, the management of acute postoperative pain is still unsatisfactory. This study was aimed to quantitatively analyze the possible correlations between inflammatory cytokines, muscle damage markers and acute postoperative pain following primary TKA. METHODS: Patients scheduled for unilateral primary TKA were consecutively included, the serial changes of the numerical rating scale (NRS) at rest (NRSR) and at walking (NRSW), serum inflammatory cytokines and muscle damage markers were assessed before surgery (T0) and at postoperative day 1, 2, 3 and 5 (T1-T4, respectively); while pain disability questionnaire (PDQ) and synovial fluid inflammatory cytokines were evaluated at T0. The correlations between inflammatory cytokines, muscle damage markers and pain scores were examined, and Bonferroni correction was applied for multiple comparisons. RESULTS: Ninety six patients were included for serum markers and pain evaluations at T0-T4, while 54 (56.25%) for synovial fluid cytokines at T0. The NRSR at T1 and T2 were positively correlated with preoperative NRSW, while the NRSW at T1 to T4 were positively correlated with preoperative NRSR, NRSW and PDQ (all p < 0.05). The NRSR was positively correlated with serum PGE2, IL-6, and CK at T1; the NRSW was positively correlated with serum CRP at T1, with PGE2 and IL-6 at T1 to T3, with CK at T2 and T4, and with Mb and LDH at T1 to T4 (all p < 0.003). Meanwhile, positive correlations were observed between preoperative NRSW and synovial fluid PGE2, IL-6, IL-8, or TNF-α, as well as between PDQ and PGE2 (all p < 0.003), but no associations between postoperative pain scores and preoperative synovial fluid cytokines was found (all p ≥ 0.003). Additionally, the NRSR at T1 and T2, and NRSW at T1 to T4 were positively correlated with body mass index (all p < 0.05). CONCLUSIONS: Serum inflammatory cytokines and muscle damage markers are positively correlated with acute postoperative pain following primary TKA, and the key cytokines (CRP, PGE2, and IL-6) and markers (Mb, CK and LDH) may serve as the targets for developing novel analgesic strategies.

[Endothelium Aging and Oxidative Stress].

In the endothelium, ROS mainly derive from mitochondria, endothelial nitric oxide synthases and NADPH oxidases 4. Excessive ROS are a major cause of oxidative stress, the primary stimulus of vascular dysfunction and oxidative stress-related diseases. However, cellular evolution has made possible the development of adaptive antioxidant systems that scavenge excessive ROS, such as Nrf2/Keapl-ARE, PPAR-y, SIRT and FOXO, etc. Among them, the Nrf2/Keapl-ARE signaling pathway is perhaps the most prominent. What is more, there are the "crosstalk" among these antioxidant stress-related signaling pathways aim to alleviate oxidative stress injurys and promote cells survival. The understanding of the relationship between endothelial aging and oxidative stress may serve as a therapeutic clues in the treatment of oxidative stress-related diseases.

Novel pyrimidin-4-one derivatives as potential T3SS inhibitors against Xanthomonas campestris pv. campestris.

BACKGROUND: Cruciferous black rot is caused by Xanthomonas campestris pv. campestris (Xcc) infection and is a widespread disease worldwide. Excessive and repeated use of bactericide is an important cause of the development of bacterial resistance. It is imperative to take new approaches to screening compounds that target virulence factors rather than kill bacterial pathogens. The type III secretion system (T3SS) invades a variety of cells by transporting virulence effector factors into the cytoplasm and is an attractive antitoxic target. Toward the search of new T3SS inhibitors, an alternative series of novel pyrimidin-4-one derivatives were designed and synthesized and assessed for their effect in blocking the virulence. RESULTS: All of the target compounds were characterized by proton (1 H) nuclear magnetic resonance (NMR), carbon-13 (13 C) NMR, fluorine-19 (19 F) NMR and high-resolution mass spectrometry (HRMS). All compounds were evaluated using high-throughput screening systems against Xcc. The results of the biological activity test revealed that the compound SPF-9 could highly inhibit the activity of xopN gene promoter and the hypersensitivity (HR) of tobacco without affecting bacterial growth. Moreover, messenger RNA (mRNA) level measurements showed that compound SPF-9 inhibited the expression of some representative genes (hrp/hrc genes). Compound SPF-9 weakened the pathogenicity of Xcc to Raphanus sativus L. CONCLUSION: Compound SPF-9 has good potential for further development as a novel T3SS inhibitor against Xcc. © 2023 Society of Chemical Industry.

[In vitro analysis and in vivo tracing of BrdU-labeled rat bone marrow mesenchymal stem cells].

OBJECTIVES: To investigate the optimal condition of bromodeoxyuridine (BrdU) labeling for bone marrow mesenchymal stem cells (BMSCs) in vitro and the feasibility of in vivo tracing of BrdU-labeling BMSCs. METHODS: BMSCs were isolated from Wistar rats and were in vitro routinely cultured. The third passage BMSCs was used for identification of special surface antigens by immunohistochemical methods. The purified BMSCs were incubated with BrdU at different concentrations for different incubating time to investigate optimal BrdU concentration and incubating time for cell labeling. The cell labeling index of BrdU was calculated with immunohistochemical analysis. BMSCs labeled BrdU were injected into damaged gastric mucosa of rats by micro injector. The colonization of BMSCs labeled BrdU in gastric mucosa was viewed. RESULTS: After purification and proliferation, the primary cultured BMSCs were uniformly long spindle-shapped form and formed cell colony, which showed the characteristics of stem cell. Immunocytochemistry showed BMSCs were positive for CD44 and CD90, while negative for CD14, CD45. The labeling rate of BrdU increased with the labeling time lasting and reached its height at 48 h. After incubating 48 and 72 hours, the labeling rate of BrdU with a concentration of 10 micromol/L was higher than that of BrdU with a concentration of 5 micromol/L (P < 0.05) and similar with that of BrdU with a concentration of 15 micromol/L (P > 0.05). In addition, the BrdU labeling could be detected after five consecutive passages and the labeling time could keep 21 d. The pathological observation demonstrated that BrdU-labeled BMSCs could colonize the damaged gastric mucosa with normal morphologic characteristics during observation period. CONCLUSION: BrdU labeling might be a feasible method for dynamic observation of the migration, growth and differentiation of migrating BMSCs in colonizing sites.

Pancreas anatomy and surgical procedure for pancreatectomy in rhesus monkeys.

BACKGROUND: The aim of this study was to investigate the pancreas anatomy and surgical procedure for harvesting pancreas for islet isolation while performing pancreatectomy to induce diabetes in rhesus monkeys. METHODS: The necropsy was performed in three cadaveric monkeys. Two monkeys underwent the total pancreatectomy and four underwent partial pancreatectomy (70-75%). RESULTS: The greater omentum without ligament to transverse colon, the cystic artery arising from the proper hepatic artery and the branches supplying the paries posterior gastricus from the splenic artery were observed. For pancreatectomy, resected pancreas can be used for islet isolation. Diabetes was not induced in the monkeys undergoing partial pancreatectomy (70-75%). CONCLUSIONS: Pancreas anatomy in rhesus monkeys is not the same as in human. Diabetes can be induced in rhesus monkeys by total but not partial pancreatectomy (70-75%). Resected pancreas can be used for islet isolation while performing pancreatectomy to induce diabetes.

[The effects of NKG2D mAb on the survival of allogeneic transplanted islets in spontaneous diabetic nonobese diabetic mice].

OBJECTIVE: To investigate the effects of NKG2D mAb on the survival of allogeneic transplanted islets nonobese diabetic (NOD) mice, and to find if CD154 mAb has synergistic effects. METHODS: Spontaneous diabetic NOD mice transplanted with allogeneic islets of BALB/c mice were divided into 4 groups. Group A was control group, Group B were treated with anti-NKG2D monoclonal antibody (mAb), Group C were treated with CD154 mAb (MR1), Group D were treated with NKG2D mAb and MR1. Glucose levels were monitored at regular intervals through caudal vein, and islet function was evaluated by glycemia. Histological study was performed at graft rejection or at day 120. Spleen cell suspension was prepared for mixed lymphocyte cultivation. The kidneys hosting the islet graft were prepared with HE staining and immuno-histochemistry staining of CD3, CD4 and CD8 was performed. RESULTS: MR1 therapy alone significantly prolonged the survival of islet grafts when compared to NKG2D mAb group and the control group: median graft survival was 41 days versus 8 days (P < 0.05) and 8 days (P < 0.05), respectively. Combination therapy with NKG2D mAb and MR1 prolonged islet grafts survival when compared to MR1 therapy alone: median graft survival was 51 days versus 41 days (P > 0.05). CONCLUSION: NKG2D mAb alone did not result in the prolongation of islet graft survival, whereas CD154 mAb increased graft survival. When both antibodies were administered, a synergistic effect was obtained, but did not provide permanent protection from diabetes recurrence.

[Toxic effect of immunosuppression agent Everolimus on insuloma cells and pancreatic islets of SD rat].

OBJECTIVE: To investigate the effect of immunosuppression agent Everolimus on the viability and function of insuloma cells (INS-1) and pancreatic islet cells cultured in vitro. METHODS: INS-1 cells and islets were treated with a series of concentrations of immunosuppression agents (Everolimus, Cyclosporin A, Sirolimus and Mycophenolate Mofetil). The viability of INS-1 cells and rat pancreatic islets were determined with MTT and the function of INS-1 cells and rat pancreatic islets was evaluated with Glucose-stimulated insulin secretion assay. RESULTS: Above the clinical blood concentration, the inhibition rate of islet cell proliferation in the high concentration group of Everolimus and Sirolimus was significantly lower than that of Cyclosporin A and Mycophenolate Mofetil group (P < 0.05); Everolimus in the blood drug level, like other immunosuppressive agents, can inhibit the function of insulin secretion, and the stimulation index of each group was no significant difference. CONCLUSION: Compared to Mycophenolate Mofetil and Cyclosporin A, Everilimus and Sirolimus demonstrate lower toxicity effect on INS-1 cells and rat pancreatic islets in vitro and Everolimus is expected as a new type of immunosuppressive agent used in clinical islet transplantation.

miR-146a-5p improves the decidual cytokine microenvironment by regulating the toll-like receptor signaling pathway in unexplained spontaneous abortion.

Spontaneous abortion (SA) is a common pregnancy failure, but the cause of numerous cases remains unexplained. Decidual immune cells (DICs)-mediated cytokine microenvironment is involved in pregnancy and regulated by many microRNAs, but whether microRNA-146a-5p (miR-146a) regulate the decidual cytokine microenvironment and the potential mechanisms in unexplained SA pathogenesis have rarely been reported. In this study, the levels of cytokines and miR-146a in healthy and unexplained SA deciduae were first investigated, and the correlation between them was analyzed. Then, the effect of miR-146a inhibitor on cytokines was assessed in healthy deciduae-derived DICs. Third, the downstream targets and related molecular mechanisms of miR-146a were analyzed by bioinformatics, and the levels of the predicted targets in deciduae were assessed, followed by the correlation analysis between the levels of miR-146a and the targets. Finally, the effect of miR-146a on the predicted targets and inflammatory cytokines was validated in unexplained SA deciduae-derived DICs. As a result, decreased miR-146a correlated with the cytokine disorder in unexplained SA deciduae, and inhibition of miR-146a promoted pro-inflammatory response in healthy deciduae-derived DICs. One hundred four target genes and related molecular mechanisms of miR-146a were predicted, among which the toll-like receptor (TLR) pathway might be associated with the decidual cytokine regulation. Upregulation of miR-146a inhibited the expression of the predicted molecules enriched in the TLR pathway and improved the cytokine disorder in unexplained SA deciduae-derived DICs. Collectively, miR-146a improves the decidual cytokine microenvironment by regulating the TLR pathway in unexplained SA, providing novel potential targets for further therapeutic research.

[Effects of enteral nutrition and parenteral nutrition on gut epithelial tight junction and barrier function in rats after surgical stress].

OBJECTIVE: To compare the effects of enteral nutrition (EN) and parenteral nutrition (PN) on gut mucosal barrier function and intestinal epithelial tight junctions in rats after surgical stress. METHODS: Fifty SD rats with surgical trauma were randomly divided into three groups: total parenteral nutrition (TPN) group and enteral nutrition (EN) group with isocaloric and isonitrogenous nutrition and placebo group. Nutrients were administered via the neck vein and needle jejunostomy for the TPN group. The homogenated tissues of liver, lung, and mesenteric lymph nodes were cultured to determine the bacterial translocations rates. The transmembrane binding proteins (occludin) were measured with immunohistochemistry. The ultrastructure and morphology of intestinal epithelial tight junctions were observed by electron microscope. The feces in cecum were cultured for anaerobic bacterial growth analyses. RESULTS: The EN group had more lactobacteria and bifydobacteria than the TPN group, but not statistical significant. The EN group had greater expression of occludin in the intestines than the TPN group. Furthermore, the intestinal epithelial tight junction and microvilli of the EN group were more intact compared with those of the TPN group. The bacterial translocations rates of liver, lung and mesenteric lymph nodes were significantly lower in the EN group than in the TPN group. CONCLUSION: Neither EN nor standard PN maintains intestinal membrane barriers. But EN increases the expression of transmember binding proteins, maintains the gut epithelial tight junction, improves intestinal mucosal barrier and reduces gut bacterial translocation.

[Effects of the different culture and isolation methods on the growth, proliferation and biology characteristics of rat bone marrow mesenchymal stem cells].

OBJECTIVE: To explore a suitable method for isolation, purification and multiplication of bone marrow mesenchymal stem cells (BMSCs). METHODS: Density gradient centrifugation and adherence separation methods were applied for isolation of BMSCs from Wistar rats. The cells were cultured and proliferated in culture medium containing calf serum (CS), fetal bovine serum (FBS), free of serum or different volume fraction of FBS. The characteristic and the morphology of BMSCs were observed under inverted microscope every day. The growth curves were draw and the surface antigen of BMSCs were detected by immunocytochemistry technique. The microstructure was observed by transmission electron microscope (TEM). RESULTS: The pure primary cells can be procured by density gradient centrifugation. But the primary cells cultured by adherence separation methods demonstrated higher cytoactive, more rapid proliferation, earlier colony confluence and shorter time for passage than that cultured by density gradient centrifugation method. The cells by adherence separation methods were essentially purified at passage 4. Both CS and FBS can promote the growth and proliferation of BMSCs, but the colony forming efficacy of cells (46.50%) cultured in medium containing 0.12 volume fraction FBS was the highest. The cells surface markers CD44, CD90 were positive and CD14, CD45 were negative. BMSCs were observed by TEM and possessed the characteristic of stem cells. Conclusion BMSCs with high quality and activity can be obtained with adherence separation by suitable method and culture conditions. L-DMEM medium containing 0.12 volume fraction FBS showed more profitable for the growth and proliferation of BMSCs.

Peripheral lung carcinoma: correlation of angiogenesis and first-pass perfusion parameters of 64-detector row CT.

PURPOSE: To investigate the value of 64-detector row CT first-pass perfusion imaging in the evaluation of tumor perfusion in patients with lung carcinoma, and to assess the correlation between the perfusion parameters and tumor angiogenesis. MATERIALS AND METHODS: Forty-six surgically peripheral lung carcinomas were examined with 64-detector row CT. First-pass CT perfusion study comprised of 12 repeated spiral acquisitions over 60s following a 50-ml intravenous bolus of contrast medium at 6-7 ml/s. Tumor specimens were assessed for microvessel density (MVD). Perfusion, peak enhancement intensity (PEI), time to peak (TTP), and blood volume (BV) and MVD of the tumor were compared by means of one-way ANOVA analysis of variance among histological type, size, metastasis and necrosis. Pearson correlation coefficients were conducted to represent the relationships between the perfusion parameters and MVD of the tumor. RESULTS: Mean values for perfusion, PEI, TTP, and BV of the 46 tumors were 70.3+/-39.4 ml/min/ml, 67.0+/-37.6 HU, 36.9+/-11.2s, and 34.9+/-17.9 ml/100g, respectively. No statistically significant differences in perfusion parameters were found among different histological types (p>0.05). Considerable differences with higher perfusion, PEI and BV were noted in tumor < or = 3.0 cm than in tumor>3.0 cm (p<0.05). No statistically significant differences were found between nodule metastasis positive and negative groups (p>0.05). The necrotic tumors showed significantly lower perfusion, PEI and BV compared with non-necrotic tumors (p<0.05). Perfusion, PEI, and BV of the necrotic part manifested significantly lower, but TTP longer, than those of non-necrotic part of the necrotic tumors (p<0.05). Perfusion, PEI and BV were positively correlated with extent of MVD (r=0.715, 0.681, 0.762, respectively, all p<0.001), whereas no significant correlation was found between TTP and MVD (r=-0.154, p>0.05). CONCLUSION: 64-detector row CT first-pass perfusion imaging is a valuable noninvasive method in evaluating tumor perfusion of peripheral lung carcinoma. CT perfusion parameters can be indicators for evaluating tumor necrosis and angiogenesis.

Expressed sequence tags analysis of a liver tissue cDNA library from a highly inbred minipig line.

BACKGROUND: Porcine liver performing efficient physiological functions in the human body is prerequisite for successful liver xenotransplantation. However, the protein differences between pig and human remain largely unexplored. Therefore, we investigated the liver expression profile of a highly inbred minipig line. METHODS: A cDNA library was constructed from liver tissue of an inbred Banna minipig. Two hundred randomly selected clones were sequenced then analysed by BLAST programme. RESULTS: Alignments of the sequences showed 44% encoded previously known porcine genes. Among the 56% unknown genes, sequences of 72 clones had high similarities with known genes of other species and the similarities to human were mostly above 0.80. The other 40 clones showing no similarity to genes in National Centre for Biotechnology Information are newly discovered, expressed sequence tags specific to liver of inbred Banna minipig. Twenty-two of the 200 clones had full length encoding regions, 38 complete 5' terminal sequences and 140 complete 3' terminal sequences. CONCLUSION: These newly discovered expression sequences may be an important resource for research involving physiological characteristics and medical usage of inbred pigs and contribute to matching studies in xenotransplantation.

[Construction of a cDNA library from liver tissue of rhesus monkey, Macaca mulatta].

OBJECTIVE: To screen the target rhesus genes and give some basic genetic evidences to its value as one of the most important animal model in biomedical study, we constructed a cDNA expression library from liver tissue of a healthy rhesus monkey. METHODS: With Trizol reagent, the total RNA was extracted from healthy rhesus liver tissue. By mutant Moloney Murine Leukemia Virus Reverse Transcriptase (MMLV-RT), the first-strand cDNA was synthesized from purified mRNA, and subsequently the second-strand cDNA was generated via E. coli DNA polymerase I . Then, the EcoR I adapter was added to the synthesized double-strand cDNA, which was subsequently digested by Xho I restriction enzyme and fractionated with CHROMA APIN-400 column. The fractionated cDNA fragments to be longer than 0. 5 kb were ligated into lambda ZAP express vector to form the phagemid cDNA recombinants, which were further packaged into the lambda ZAP cDNA library according to the standard protocol with phage lambda Gold packaging extract. In order to get more stable clones with larger quantity, the primary library was amplified through infecting the host strain XL1-Blue MRF'. Then, the library titre, recombinant rate and length of inserted cDNA were measured, respectively. RESULTS: The capacity of the primary stand or unamplified library was 1. 2X 10(6) pfu. The titers of the unamplified library or the amplified library was 1.1 X 10(6) mixture, pfu/mL or 7. 7 X 10(9) pfu/mL respectively, the percentages of recombinants were 99. 3% and 98. 2%, and the average lengths of the inserts were 2.0 kb and 2. 3 kb, respectively. CONCLUSION: An excellent cDNA expression library has been constructed successfully, which would lay solid foundation for transplantation study and pre-clinic evaluation of related drugs.

[Interaction between member LIGHT of TNF superfamily and SOCS3, which respond to induce the differentiation and maturation of dendritic cell].

OBJECTIVE: To detect the relation between the member LIGHT of TNF superfamily and the suppressor of cytokine signaling 3 (SOCS3), and to investigate the effect of SOCS3 on dendritic cell (DC) maturation induced by LIGHT. METHODS: Bone marrow-derived DC (BMDC) was generated from mouse bone marrow monocyte by culturing with rmGM-CSF, rmIL-4 in vitro. SOCS3 mRNA in BMDC was analyzed by RT-PCR, and the protein of SOCS3 was measured by Western blot. After blocking the SOCS3 expression with the specific anti-sense oligonucleotide, we applied the flow cytometry to measure the expression of CD86 and CD40 on DC for making clear whether the silence of SOCS3 would regulate the LIGHT-stimulated DC maturation. RESULTS: With the effect of LIGHT, the level of SOCS3 mRNA and protein in BMDC sharply increased. The specific antisense oligonucleotide could effectively block SOCS3 mRNA expressing in BMDC with the ratio of 49% and block SOCS3 protein expression with the ratio of 45%. Compared with SOCS3-unblocked DC, the SOCS3-blocked BMDC with stimulation of LIGHT showed higher CD40 and CD86 (P < 0.05). CONCLUSION: LIGHT enhances the expression of SOCS3 during stimulating BMDC maturation. As more sensitive to LIGHT, the SOCS3-blocked BMDC is driven to more mature. SOCS3 presents a negative regulation mechanism in BMDC maturation induced

[Simultaneous pancreas-kidney transplantation (SPK) for an old age male with type 2 diabetes complicated with end-stage renal disease (ESRD): a case report].

We reported the first case of simultaneous pancreas-kidney transplantation (SPK) in our hospital. The recipient is a 65 year old male, who suffered type 2 diabetes for 15 years and renal dysfunction for 5 years and other diabetic complications such as retinopathy, peripheral neuropathy. SPK was performed successfully for him in March, 2007, in which the donor kidney was put in left iliac fossa, while the donor pancreas grafted to set in right iliac fossa of recipient, with pancreas exocrine drainage controlled by anastomosis to the small bowel and endocrine release done to the circulatory system. Serum C-peptide, Creatinine and Blood urea nitrogen became normal levels at day 1, 4 and 11 of post-operation respectively. The concentration of blood glucose was stabilized gradually to normal level and therefore the injected insulin was stopped using to the patient at day 16 of post-operative days. OGTT test showed the function of grafted pancreas was normal 3 weeks after transplant, and no transplantation-related complications occurred. With the recipient followed up for 6 months, both his blood glucose level and renal function maintained normal without using injected insulin, and he was getting to recover from other diabetic complications also.

Control-released basic fibroblast growth factor-loaded poly-lactic-co-glycolic acid microspheres promote sciatic nerve regeneration in rats.

Although peripheral nerve injury may result in a loss of function in innervated areas, the most effective method for nerve regeneration remains to be determined. The aim of the present study was to investigate the effect of control-released basic fibroblast growth factor (bFGF)-loaded poly-lactic-co-glycolic acid (PLGA) microspheres on sciatic nerve regeneration following injury in rats. bFGF-PLGA microspheres were prepared and their characteristics were evaluated. The sciatic nerve was segmentally resected to create a 10 mm defect in 36 Sprague Dawley (SD) rats and, following the anastomosis of the nerve ends with a silicone tube, bFGF-PLGA microspheres, free bFGF or PBS were injected into the tube (n=12 in each group). The outcome of nerve regeneration was evaluated using the sciatic function index (SFI), electrophysiological test and histological staining at 6 weeks and 12 weeks post-surgery. The bFGF-PLGA microspheres were successfully synthesized with an encapsulation efficiency of 66.43%. The recovery of SFI and electrophysiological values were significantly greater (P<0.05), and morphological and histological observations were significantly greater (P<0.05) in bFGF-PLGA microspheres and bFGF groups compared with those in the PBS group, and the quickest recovery was observed in the bFGF-PLGA microspheres group. In conclusion, the bFGF-PLGA microspheres may promote nerve regeneration and functional recovery in the sciatic nerve, and may have potential therapeutic applications in peripheral nerve regeneration.

The effect of primary total knee arthroplasty on the incidence of falls and balance-related functions in patients with osteoarthritis.

Knee osteoarthritis (OA) is an established risk factor for falls and balance impairment. This study investigated the incidence of falls, balance-related outcomes and risk factors for falls before and after primary total knee arthroplasty (TKA). Three hundred seventy-six OA patients scheduled to undergo TKA were included. Falls data within the preoperative, first postoperative and second postoperative years were collected, balance-related functions were assessed using the Assessment of Quality of Life (AQoL), WOMAC, Falls Efficacy Scale International (FES-I), Activities-specific Balance Confidence (ABC), knee extension strength, Berg Balance Scale (BBS) and Timed Up and Go (TUG) before surgery and 1 and 2 years after surgery. Compared with preoperative values, the incidence of falls significantly decreased (14.89%, 6.23% and 3.14% within the preoperative, first postoperative and second postoperative years, respectively) and the AQoL, WOMAC, FES-I, ABC, knee extension strength, BBS and TUG significantly improved after TKA. Logistic regression analysis revealed that Kellgren-Lawrence grade ≥ 3 of the contralateral knee was an independent risk factor for falls before and after TKA. Conclusively, primary TKA is associated with a reduced incidence of falls and improved balance-related functions, and the contralateral knee should be considered in the design of fall-prevention strategies in patients with OA.

[HPLC determination and diversity analysis of thiopurine methyltransferase activity in human and pig].

OBJECTIVE: To develop an HPLC method for the determination of 6-methylmercaptopurine concentration and then numerate the thiopurine methyltransferase (TPMT) activity in erythrocyte via formula and analysis of the TPMT activity difference between human and pig. METHODS: The chromatographic apparatus SHIMADZU GC-2010C was used. The stationary phase was a Symmetry C18 reverse-phase column (150 mm x 3.9 mm, 5 microm). The mobile phase was 0.1% acetic acid-methanol and the flow rate 1.0 mL/min. The samples were extracted by ethyl acetate and injected automatically. They were measured at UV 280 nm. 2-amino-6-methyl-mercaptopurine was used as the internal standard. RESULTS: The retention times for 6-MP, AMMP and 6-MMP were 4.4 min, 7.7 min and 9.3 min respectively. The calibration curves were linear over the range of 6.25-100 nmol/mL. the methodology recovery was 98.08%-100.05%. The extraction recovery of 6-MMP was 88.1%-92.4%. The within-group RSD 1.0%-1.5% and inter-group RSD 1.0%-4.4%. The levels of TPMT activity of human, pig and Banna Minipig Inbred Line (BMI) were (17.45 +/- 3.62) U, (7.65 +/- 1.35) U and (8.73 +/- 1.55) U respectively. CONCLUSION: The method is suitable for determination of TPMT activity in erythrocyte of different patients and species in that of clinical medicine and scientific research. TPMT activity of pig and BMI is obviously lower than human's.

Growth inhibition of high-intensity focused ultrasound on hepatic cancer in vivo.

AIM: To investigate the damaging effect of high-intensity focused ultrasound (HIFU) on cancer cells and the inhibitory effect on tumor growth. METHODS: Murine H22 hepatic cancer cells were treated with HIFU at the same intensity for different lengths of time and at different intensities for the same length of time in vitro, the dead cancer cells were determined by trypan blue staining. Two groups of cancer cells treated with HIFU at the lowest and highest intensity were inoculated into mice. Tumor masses were removed and weighed after 2 wk, tumor growth in each group was confirmed pathologically. RESULTS: The death rate of cancer cells treated with HIFU at 1 000 W/cm2 for 0.5, 1, 2, 4, 8, and 12 s was 3.11+/-1.21%, 13.37+/-2.56%, 38.84+/-3.68%, 47.22+/-5.76%, 87.55+/-7.32%, and 94.33+/-8.11%, respectively. A positive relationship between the death rates of cancer cells and the length of HIFU treatment time was found (r = 0.96, P<0.01). The death rate of cancer cells treated with HIFU at the intensity of 100, 200, 400, 600, 800, and 1 000 W/cm2 for 8 s was 26.31+/-3.26%, 31.00+/-3.87%, 41.97+/-5.86%, 72.23+/-8.12%, 94.90+/-8.67%, and 99.30+/-9.18%, respectively. A positive relationship between the death rates of cancer cells and the intensities of HIFU treatment was confirmed (r = 0.98, P<0.01). The cancer cells treated with HIFU at 1 000 W/cm2 for 8 s were inoculated into mice ex vivo. The tumor inhibitory rate was 90.35% compared to the control (P<0.01). In the experimental group inoculated with the cancer cells treated with HIFU at 1 000 W/cm2 for 0.5 s, the tumor inhibitory rate was 22.9% (P<0.01). By pathological examination, tumor growth was confirmed in 8 out of 14 mice (57.14%, 8/14) inoculated with the cancer cells treated with HIFU at 1 000 W/cm2 for 8 s, which was significantly lower than that in the control (100%, 15/15, P<0.05). CONCLUSION: HIFU is effective on killing or damage of H22 hepatic cancer cells in vitro and on inhibiting tumor growth in mice ex vivo.