Mitochondrial fumarate promotes ischemia/reperfusion-induced tubular injury.

AIM: Mitochondrial dysfunction, a characteristic pathological feature of renal Ischemic/reperfusion injury (I/RI), predisposes tubular epithelial cells to maintain an inflammatory microenvironment, however, the exact mechanisms through which mitochondrial dysfunction modulates the induction of tubular injury remains incompletely understood. METHODS: ESI-QTRAP-MS/MS approach was used to characterize the targeted metabolic profiling of kidney with I/RI. Tubule injury, mitochondrial dysfunction, and fumarate level were evaluated using qPCR, transmission electron microscopy, ELISA, and immunohistochemistry. RESULTS: We demonstrated that tubule injury occurred at the phase of reperfusion in murine model of I/RI. Meanwhile, enhanced glycolysis and mitochondrial dysfunction were found to be associated with tubule injury. Further, we found that tubular fumarate, which resulted from fumarate hydratase deficiency and released from dysfunctional mitochondria, promoted tubular injury. Mechanistically, fumarate induced tubular injury by causing disturbance of glutathione (GSH) hemostasis. Suppression of GSH with buthionine sulphoximine administration could deteriorate the fumarate inhibition-mediated tubule injury recovery. Reactive oxygen species/NF-κB signaling activation played a vital role in fumarate-mediated tubule injury. CONCLUSION: Our studies demonstrated that the mitochondrial-derived fumarate promotes tubular epithelial cell injury in renal I/RI. Blockade of fumarate-mediated ROS/NF-κB signaling activation may serve as a novel therapeutic approach to ameliorate hypoxic tubule injury.

Activation of HIF-1α C-terminal transactivation domain protects against hypoxia-induced kidney injury through hexokinase 2-mediated mitophagy.

The transcription factor hypoxia-inducible factor-1α (HIF-1α), as a master regulator of adaptive responses to hypoxia, possesses two transcriptional activation domains [TAD, N-terminal (NTAD), and C-terminal (CTAD)]. Although the roles of HIF-1α NTAD in kidney diseases have been recognized, the exact effects of HIF-1α CTAD in kidney diseases are poorly understood. Here, two independent mouse models of hypoxia-induced kidney injury were established using HIF-1α CTAD knockout (HIF-1α CTAD-/-) mice. Furthermore, hexokinase 2 (HK2) and mitophagy pathway are modulated using genetic and pharmacological methods, respectively. We demonstrated that HIF-1α CTAD-/- aggravated kidney injury in two independent mouse models of hypoxia-induced kidney injury, including ischemia/reperfusion-induced kidney injury and unilateral ureteral obstruction-induced nephropathy. Mechanistically, we found that HIF-1α CTAD could transcriptionally regulate HK2 and subsequently ameliorate hypoxia-induced tubule injury. Furthermore, it was found that HK2 deficiency contributed to severe renal injury through mitophagy inhibition, while mitophagy activation using urolithin A could significantly protect against hypoxia-induced kidney injury in HIF-1α C-TAD-/- mice. Our findings suggested that the HIF-1α CTAD-HK2 pathway represents a novel mechanism of kidney response to hypoxia, which provides a promising therapeutic strategy for hypoxia-induced kidney injury.

Disturbance of Hypoxia Response and Its Implications in Kidney Diseases.

Significance: The disturbance of the hypoxia response system is closely related to human diseases, because it is essential for the maintenance of homeostasis. Given the significant role of the hypoxia response system in human health, therapeutic applications targeting prolyl hydroxylase-hypoxia-inducible factor (HIF) signaling have been attempted. Thus, systemically reviewing the hypoxia response-based therapeutic strategies is of great significance. Recent Advances: Disturbance of the hypoxia response is a characteristic feature of various diseases. Targeting the hypoxia response system is, thus, a promising therapeutic strategy. Interestingly, several compounds and drugs are currently under clinical trials, and some have already been approved for use in the treatment of certain human diseases. Critical Issues: We summarize the molecular mechanisms of the hypoxia response system and address the potential therapeutic implications in kidney diseases. Given that the effects of hypoxia response in kidney diseases are likely to depend on the pathological context, specific cell types, and the differences in the activation pattern of HIF isoforms, the precise application is critical for the treatment of kidney diseases. Although HIF-PHIs (HIF-PHD inhibitors) have been proven to be effective and well tolerated in chronic kidney disease patients with anemia, the potential on-target consequence of HIF activation and some outstanding questions warrant further consideration. Future Direction: The mechanism of the hypoxia response system disturbance remains unclear. Elucidation of the molecular mechanism of hypoxia response and its precise effects on kidney diseases warrants clarification. Considering the complexity of the hypoxia response system and multiple biological processes controlled by HIF signaling, the development of more specific inhibitors is highly warranted. Antioxid. Redox Signal. 37, 936-955.

SAP130 released by damaged tubule drives necroinflammation via miRNA-219c/Mincle signaling in acute kidney injury.

Tubules injury and immune cell activation are the common pathogenic mechanisms in acute kidney injury (AKI). However, the exact modes of immune cell activation following tubule damage are not fully understood. Here we uncovered that the release of cytoplasmic spliceosome associated protein 130 (SAP130) from the damaged tubular cells mediated necroinflammation by triggering macrophage activation via miRNA-219c(miR-219c)/Mincle-dependent mechanism in unilateral ureteral obstruction (UUO) and cisplatin-induced AKI mouse models, and in patients with acute tubule necrosis (ATN). In the AKI kidneys, we found that Mincle expression was tightly correlated to the necrotic tubular epithelial cells (TECs) with higher expression of SAP130, a damaged associated molecule pattern (DAMP), suggesting that SAP130 released from damaged tubular cells may trigger macrophage activation and necroinflammation. This was confirmed in vivo in which administration of SAP130-rich supernatant from dead TECs or recombinant SAP130 promoted Mincle expression and macrophage accumulation which became worsen with profound tubulointerstitial inflammation in LPS-primed Mincle WT mice but not in Mincle deficient mice. Further studies identified that Mincle was negatively regulated via miR-219c-3p in macrophages as miR-219c-3p bound Mincle 3'-UTR to inhibit Mincle translation. Besides, lentivirus-mediated renal miR-219c-3p overexpression blunted Mincle and proinflammatory cytokine expression as well as macrophage infiltration in the inflamed kidney of UUO mice. In conclusion, SAP130 is released by damaged tubules which elicit Mincle activation on macrophages and renal necroinflammation via the miR-219c-3p-dependent mechanism. Results from this study suggest that targeting miR-219c-3p/Mincle signaling may represent a novel therapy for AKI.

Kim-1 Targeted Extracellular Vesicles: A New Therapeutic Platform for RNAi to Treat AKI.

BACKGROUND: AKI is a significant public health problem with high morbidity and mortality. Unfortunately, no definitive treatment is available for AKI. RNA interference (RNAi) provides a new and potent method for gene therapy to tackle this issue. METHODS: We engineered red blood cell-derived extracellular vesicles (REVs) with targeting peptides and therapeutic siRNAs to treat experimental AKI in a mouse model after renal ischemia/reperfusion (I/R) injury and unilateral ureteral obstruction (UUO). Phage display identified peptides that bind to the kidney injury molecule-1 (Kim-1). RNA-sequencing (RNA-seq) characterized the transcriptome of ischemic kidney to explore potential therapeutic targets. RESULTS: REVs targeted with Kim-1-binding LTH peptide (REVLTH) efficiently homed to and accumulated at the injured tubules in kidney after I/R injury. We identified transcription factors P65 and Snai1 that drive inflammation and fibrosis as potential therapeutic targets. Taking advantage of the established REVLTH, siRNAs targeting P65 and Snai1 were efficiently delivered to ischemic kidney and consequently blocked the expression of P-p65 and Snai1 in tubules. Moreover, dual suppression of P65 and Snai1 significantly improved I/R- and UUO-induced kidney injury by alleviating tubulointerstitial inflammation and fibrosis, and potently abrogated the transition to CKD. CONCLUSIONS: A red blood cell-derived extracellular vesicle platform targeted Kim-1 in acutely injured mouse kidney and delivered siRNAs for transcription factors P65 and Snai1, alleviating inflammation and fibrosis in the tubules.

Exosomal miR-125b-5p deriving from mesenchymal stem cells promotes tubular repair by suppression of p53 in ischemic acute kidney injury.

Mesenchymal stem cells-derived exosomes (MSC-exos) have attracted great interest as a cell-free therapy for acute kidney injury (AKI). However, the in vivo biodistribution of MSC-exos in ischemic AKI has not been established. The potential of MSC-exos in promoting tubular repair and the underlying mechanisms remain largely unknown. Methods: Transmission electron microscopy, nanoparticle tracking analysis, and western blotting were used to characterize the properties of human umbilical cord mesenchymal stem cells (hucMSCs) derived exosomes. The biodistribution of MSC-exos in murine ischemia/reperfusion (I/R) induced AKI was imaged by the IVIS spectrum imaging system. The therapeutic efficacy of MSC-exos was investigated in renal I/R injury. The cell cycle arrest, proliferation and apoptosis of tubular epithelial cells (TECs) were evaluated in vivo and in HK-2 cells. The exosomal miRNAs of MSC-exos were profiled by high-throughput miRNA sequencing. One of the most enriched miRNA in MSC-exos was knockdown by transfecting miRNA inhibitor to hucMSCs. Then we investigated whether this candidate miRNA was involved in MSC-exos-mediated tubular repair. Results:Ex vivo imaging showed that MSC-exos was efficiently homing to the ischemic kidney and predominantly accumulated in proximal tubules by virtue of the VLA-4 and LFA-1 on MSC-exos surface. MSC-exos alleviated murine ischemic AKI and decreased the renal tubules injury in a dose-dependent manner. Furthermore, MSC-exos significantly attenuated the cell cycle arrest and apoptosis of TECs both in vivo and in vitro. Mechanistically, miR-125b-5p, which was highly enriched in MSC-exos, repressed the protein expression of p53 in TECs, leading to not only the up-regulation of CDK1 and Cyclin B1 to rescue G2/M arrest, but also the modulation of Bcl-2 and Bax to inhibit TEC apoptosis. Finally, inhibiting miR-125b-5p could mitigate the protective effects of MSC-exos in I/R mice. Conclusion: MSC-exos exhibit preferential tropism to injured kidney and localize to proximal tubules in ischemic AKI. We demonstrate that MSC-exos ameliorate ischemic AKI and promote tubular repair by targeting the cell cycle arrest and apoptosis of TECs through miR-125b-5p/p53 pathway. This study provides a novel insight into the role of MSC-exos in renal tubule repair and highlights the potential of MSC-exos as a promising therapeutic strategy for AKI.

Bone marrow mesenchymal stem cells stimulated with low-intensity pulsed ultrasound: Better choice of transplantation treatment for spinal cord injury: Treatment for SCI by LIPUS-BMSCs transplantation.

Stem cell transplantation, especially treatment with bone marrow mesenchymal stem cells (BMSCs), has been considered a promising therapy for the locomotor and neurological recovery of spinal cord injury (SCI) patients. However, the clinical benefits of BMSCs transplantation remain limited because of the considerably low viability and inhibitory microenvironment. In our research, low-intensity pulsed ultrasound (LIPUS), which has been widely applied to clinical applications and fundamental research, was employed to improve the properties of BMSCs. The most suitable intensity of LIPUS stimulation was determined. Furthermore, the optimized BMSCs were transplanted into the epicenter of injured spinal cord in rats, which were randomized into four groups: (a) Sham group (n = 10), rats received laminectomy only and the spinal cord remained intact. (b) Injury group (n = 10), rats with contused spinal cord subjected to the microinjection of PBS solution. (c) BMSCs transplantation group (n = 10), rats with contused spinal cord were injected with BMSCs without any priming. (d) LIPUS-BMSCs transplantation group (n = 10), BMSCs stimulated with LIPUS were injected at the injured epicenter after contusion. Rats were then subjected to behavioral tests, immunohistochemistry, and histological observation. It was found that BMSCs stimulated with LIPUS obtained higher cell viability, migration, and neurotrophic factors expression in vitro. The rate of apoptosis remained constant. After transplantation of BMSCs and LIPUS-BMSCs postinjury, locomotor function was significantly improved in LIPUS-BMSCs transplantation group with higher level of brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) in the epicenter, and the expression of neurotrophic receptor was also enhanced. Histological observation demonstrated reduced cavity formation in LIPUS-BMSCs transplantation group when comparing with other groups. The results suggested LIPUS can improve BMSCs viability and neurotrophic factors expression in vitro, and transplantation of LIPUS-BMSCs could promote better functional recovery, indicating possible clinical application for the treatment of SCI.

Nafamostat mesilate attenuates inflammation and apoptosis and promotes locomotor recovery after spinal cord injury.

AIM: Spinal cord injury (SCI) leads to severe neural damage for which there is currently no effective treatment. Exploration of the neuroprotective effect among clinically approved drugs will speed up clinical translation of SCI. Nafamostat mesilate (NM) as a synthetic serine protease inhibitor has been used clinically in pancreatitis treatments. However, its effectiveness in SCI is unknown. The aim of this study was to confirm the efficacy of NM in ameliorating SCI. METHODS: Intraperitoneal administration of NM was performed on a contusion SCI model in Wistar rat. Hematoxylin and eosin staining (H&E staining) and Luxol fast blue (LFB) staining were used to observe the histological lesions. Apoptosis was examined by TUNEL staining, Annexin V-FITC/PI, caspase-3, and Bcl-2. Cytokines and neurotrophins were tested by Western blot. Locomotion recovery assessed by hindlimb BBB score and the inclined plane test. RESULTS: Nafamostat mesilate treatment significantly improved locomotion recovery as assessed by hindlimb BBB scores and the inclined plane test. H&E staining and LFB staining showed a significant increase in spared tissue in both gray matter and white matter. NM decreased the expression of the proinflammatory cytokines TNF-α and IL-6. In addition, apoptosis was also significantly decreased, as shown by TUNEL staining and Annexin V-FITC/PI and by Western blotting for caspase-3 and Bcl-2 expression. Due to the mechanism of action of NM as a serine protease inhibitor, the drug decreased thrombin expression in the damaged spinal cord. Furthermore, NM increased the expression of neurotrophins (NT-3, BDNF, and NGF). CONCLUSIONS: Upon NM treatment, the functional and histological outcomes were improved, and microenvironment upon SCI was modulated. As a clinically approved drug, NM holds promise for clinical use after spinal cord injury.

The carcinogenicity of alendronate in patients with osteoporosis: evidence from cohort studies.

CONTEXT: Alendronate may relate to the incidence of cancers, especially esophageal and colon cancer. But the results are inconsistent in different studies. OBJECTIVE: To quantify the association between the use of alendronate and the occurrence of different types of cancer. DATA SOURCES: We searched Embase, Pubmed, CENTRAL, SIGLE and clinicaltrials.gov, up to 2014 June. STUDY SELECTION: Cohort studies reporting association between alendronate or bisphosphonate therapy including alendronate in patients with osteoporosis and risk of cancer were selected by two authors. DATA EXTRACTION: Two authors independently extracted the data. The Chi-square test and the I-square test were used for testing heterogeneity between studies. DATA SYNTHESIS: Eight cohort studies were included in the meta-analysis. Meta-analysis result manifested that alendronate significantly increased the incidence of lung cancer (HR 1.23, 95%CI 1.03 to 1.47, P value = 0.03), nevertheless, there was no significant difference after we excluded either Lee's 2012 study (HR 1.17, 95%CI 0.95 to 1.44, P value = 0.13) or Chiang's 2012 study (HR 1.47, 95%CI 1 to 2.17, P value = 0.05). For the incidence of colorectal cancer, no significant difference occurred (HR 0.91, 95%CI 0.74 to 1.13, P value = 0.39), but there was a positive relationship when we used fixed model (HR 0.85, 95%CI 0.78 to 0.93, P value = 0.004). For the incidence of liver cancer, there was no significant difference (HR 1.36, 95%CI 0.9 to 2.04, P value = 0.14), however, the result changed after we excluded Chiang's 2012 study (HR 1.69, 95%CI 1.03 to 2.77, P value = 0.04). There was no significant difference in other types of cancer. CONCLUSION: Based on current evidences, alendronate therapy may be associated with a high risk of lung cancer, may with an excess risk of liver cancer, a low risk of colorectal and no related risk of other cancers.

Comparative efficacy and tolerability of three treatments in old people with osteoporotic vertebral compression fracture: a network meta-analysis and systematic review.

PURPOSE: The question which kind of methods is most suitable for treating the old people for osteoporotic vertebral compression fracture is still discussed and pairwise meta-analyses cannot get hierarchies of these treatments. Our aim is to integrate the evidence to provide hierarchies of the comparative efficacy measured by the change of VAS (Visual Analogue Scale) and tolerability measured by incidence of new fractures and risk of all-cause discontinuation on three treatments (percutaneous vertebroplasty (PVP)、balloon kyphoplasty (BK) and conservative treatment (CT)). METHODS: We performed a Bayesian-framework network meta-analysis of randomized controlled trials (RCTs) to compare three treatments for the old people with osteoporotic vertebral compression fracture. The eligible RCTs were identified by searching Amed, British Nursing Index, Embase, Pubmed, the Cochrane Central Register of Controlled Trials (CENTRAL), Google scholar, SIGLE, the National Technical Information Service, the National Research Register (UK) and the Current Controlled Trials databases. Data from three outcomes (e.g. VAS, risk of all-cause discontinuation and incidence of new fractures) were independently extracted by two authors. RESULTS: A total of five RCTs were finally included into this article. PVP and BK significantly decreased VAS when compared with CT. BK had a significantly lower risk of all-cause discontinuation contrast to CT. Three treatments (BK, PVP and CT) had no significant differences in the incidence of new fractures. CONCLUSIONS: PVP may be the best way to relieve pain, CT might lead to the lowest incidence of new fractures and BK might had the lowest risk of all-cause discontinuation in old people with osteoporotic vertebral compression fracture. More large-scale and longer duration of follow-up studies are needed.

Evidence-based computer-navigated total hip arthroplasty: an updated analysis of randomized controlled trials.

BACKGROUND: Total hip arthroplasty (THA) has evolved over the years to be a reliable, reproducible, and successful orthopedic procedure. Nowadays, THA is increasingly performed on patients using less invasive, tissue-preserving techniques. Accordingly, the use of computer navigation in total joint arthroplasty has become more prevalent. However, there is still lack of high-quality evidence to verify the most effective technique for THA. METHODS: A search was conducted in PubMed, Medline, Embase, Cochrane Central Register of Controlled Trials, and Google Scholar databases. Clinical trials published from 1966 to Feb 2012 that assess conventional techniques THA or computer-navigated techniques THA for placing the acetabular component. The main outcome measures included abduction angles, anteversion angles, percentage of acetabular outliers, operation time, decrease in Hb/24 h, and wound secretion/48 h. RESULTS: The pooled analysis across all studies showed a significant difference in anteversion angles and acetabular outliers (difference -0.22, 95% CI -0.67, 0.24; p = 0.346, I (2) = 71.9%) and (difference 8.34, 95% CI 4.15, 16.74; p = 0.000, I (2) = 0.0%). However, no significant difference in abduction angle and decrease in Hb/24 h (difference -0.22, 95% CI -0.67, 0.24; p = 0.346, I (2) = 71.9%) and (difference 0.03, 95% CI -0.36, 0.41; p = 0.888, I (2) = 0.0%). For the operation time, computer-navigated THA was longer (difference -0.73, 95% CI -1.32, -0.15; p = 0.014, I (2) = 74.4%). CONCLUSIONS: This meta-analysis demonstrated computer-navigated THA was a more favorable method for placing the acetabular component and decreased the number of acetabular cups implanted outside the desired range of alignment. More high-quality RCTs were needed to support the evidence.

Tourniquet used in anterior cruciate ligament reconstruction: a system review.

PURPOSE: To identify whether routine use of a tourniquet is a better choice for anterior cruciate ligament reconstruction. METHOD: We searched Amed, British Nursing Index, Embase, Pubmed, Scopus, Cochrane Library and Google Scholar. We used revised Jadad score to evaluate the trial quality. Each reference list was viewed for any ignored studies. Two reviews independently extracted data from all eligible trials, including study design, patients' characteristics, interventions and outcomes. The available data were using random effects models with mean differences for continuous variables. RESULTS: The only meta-analysis indicated there was no significant difference in operative time between the tourniquet and non-tourniquet groups (mean differences -5.71, 95 % CI -12.40, 0.99). The remaining outcomes had variations in the outcome measures, so it was not possible to perform meta-analysis. CONCLUSIONS: There was insufficient evidence to support the hypothesis that patients would benefit from routinely applying a tourniquet. More high-quality randomized controlled trials were needed to test the result.

Cemented versus uncemented hemiarthroplasty for displaced femoral neck fractures: an updated meta-analysis.

OBJECTIVES: To compare the outcomes of cemented and uncemented hemiarthroplasty for treating displaced femoral neck fractures. METHOD: We searched the PubMed, Medline, Embase, Cochrane Central Register of Controlled Trials, and Google Scholar databases from 1966 to Mar 2012. No language restriction was applied. Reference lists of all the selected articles were hand-searched for any additional trials. Trial quality was assessed using the modified Jadad Scale. Two authors independently extracted data from all eligible studies, including study design, participants, interventions, and outcomes (mortality, hospital stay, blood loss, operation time, residual pain, and complications). The data were using fixed-effects and random-effects models with mean differences and risk ratios for continuous and dichotomous variables, respectively. RESULTS: A total of 12 studies involving 1805 patients were identified in this analysis. Meta-analysis showed longer operation time (SMD, -0.43, 95 % CI -0.56, -0.30) in cemented versus uncemented hemiarthroplasty. There was no significant difference between the two treatment groups regarding mortality (OR, 1.08, 95 % CI 0.88, 1.34), hospital stay (SMD, -1.21, 95 % CI -2.24, -0.18), blood loss (SMD, -0.12, 95 % CI -0.33, 0.10), operation time (SMD, -0.43, 95 % CI -0.56, -0.30), residual pain (OR, 1.42, 95 % CI 0.99, 2.03), and complications (OR, 0.82, 95 % CI 0.63, 1.08). CONCLUSIONS: The available evidence suggested there was no significant difference between uncemented and cemented hemiarthroplasty in treating displaced femoral neck fractures.

Single-bundle or double-bundle for anterior cruciate ligament reconstruction: a meta-analysis.

PURPOSE: To compare the clinical outcomes of anterior cruciate ligament (ACL) reconstruction with double-bundle and single-bundle techniques. STUDY DESIGN: Meta-analysis METHODS: We searched electronic databases including PubMed, Embase, Cochrane Central Register of Controlled Trials, and Google Scholar from 1966 to Jan 2012 to identify randomized controlled trials (RCTs) comparing clinical outcomes of anterior cruciate ligament (ACL) reconstruction with double-bundle and single-bundle techniques. Two reviewers independently extracted data and assessed trial quality. Meta-analysis was performed to pool results. RESULTS: Nineteen RCTs were included with a total of 1686 patients. The pooled analysis across all studies showed that the double-bundle ACL reconstruction technique could have significantly better outcomes in rotational laxity, as assessed by the pivot-shift test, KT grading and IKDC grading than the single-bundle techniques. We found no evidence of a difference in function measured by IKDC scores, KT arthrometer, Lysholm knee, or Tegner activity scores and complications after operations between single and double-bundle ACL reconstruction groups. CONCLUSION: Our meta-analysis demonstrated the superiority of double-bundle over single-bundle anterior cruciate ligament reconstruction. The double-bundle ACL reconstruction technique has better outcomes in rotational laxity (pivot-shift test, KT grading and IKDC grading). However, for functional recovery, there was no significant difference between single-bundle and double-bundle reconstruction techniques.

No difference in clinical outcome between fixed- and mobile-bearing TKA: a meta-analysis.

PURPOSE: To compare the clinical and radiographic results of fixed-bearing and mobile-bearing total knee arthroplasty (TKA). METHODS: We searched the PubMed, Medline, Embase, Cochrane Central Register of Controlled Trials, and Google Scholar databases from 1966 to January 2012. No language restriction was applied. Reference lists of all the selected articles were hand-searched for any additional trials. Trial quality was assessed using the modified Jadad scale. Two authors independently extracted data from all eligible studies, including study design, participants, interventions, and outcomes (Knee Society Score, range of movement, radiolucent line, patient preference, walking support, pain score, and complications). The data were using fixed-effects or random-effects models with mean differences and risk ratios for continuous and dichotomous variables, respectively. RESULTS: A total of 24 studies involving 2,799 patients were identified in this analysis. Meta-analysis showed lower pain score (OR, 0.66, 95% CI 0.46, 0.94) in mobile-bearing TKA than fixed-bearing TKA. There was no significant difference between the two treatment groups regarding Knee Society Score (SMD, -0.17, 95% CI: -0.60, 0.26), range of movement (SMD, -0.05, 95% CI: -0.63, 0.53), radiolucent line (OR, 1.03, 95% CI 0.74, 1.44), patient preference (OR, 1.15, 95% CI 0.82, 1.61), walking support (OR, 1.07, 95% CI 0.68, 1.70), and complications (OR, 0.85, 95% CI 0.59, 1.21). CONCLUSIONS: The available evidence suggested that there was no significant difference between clinical and radiographic results of fixed-bearing and mobile-bearing TKA except for pain score. Regarding clinical relevance, the less incidence of pain could be the advantage for selecting mobile-bearing TKA. LEVEL OF EVIDENCE: II.

UNC-64 and RIC-4, the plasma membrane-associated SNAREs syntaxin and SNAP-25, regulate fat storage in nematode Caenorhabditis elegans.

OBJECTIVE: To investigate whether genes required for synaptogenesis and synaptic function are also involved in fat storage control in Caenorhabditis elegans. METHODS: Fat storage was examined in mutants of genes affecting the synaptogenesis and synaptic function. In addition, the genetic interactions of SNAREs syntaxin/unc-64 and SNAP-25/ric-4 with daf-2, daf-7, nhr-49, sbp-1 and mdt-15 in regulating fat storage were further investigated. The tissue-specific activities of unc-64 and ric-4 were investigated to study the roles of unc-64 and ric-4 in regulating fat storage in the nervous system and/or the intestine. RESULTS: Mutations of genes required for the formation of presynaptic neurotransmission site did not obviously influence fat storage. However, among the genes required for synaptic function, the plasma membrane-associated SNAREs syntaxin/unc-64 and SNAP-25/ric-4 genes were involved in the fat storage control. Fat storage in the intestinal cells was dramatically increased in unc-64 and ric-4 mutants as revealed by Sudan Black and Nile Red strainings, although the fat droplet size was not significantly changed. Moreover, in both the nervous system and the intestine, expression of unc-64 significantly inhibited the increase in fat storage observed in unc-64 mutant. And expression of ric-4 in the nervous system completely restored fat storage in ric-4 mutant. Genetic interaction assay further indicated that both unc-64 and ric-4 regulated fat storage independently of daf-2 [encoding an insulin-like growth factor-I (IGF-I) receptor], daf-7 [encoding a transforming growth factor-beta (TGF-beta) ligand], and nhr-49 (encoding a nuclear hormone receptor). Besides, mutation of daf-16 did not obviously affect the phenotype of increased fat storage in unc-64 or ric-4 mutant. Furthermore, unc-64 and ric-4 regulated fat storage probably through the ARC105/mdt-15- and SREBP/sbp-1-mediated signaling pathways. In addition, fat storage in unc-64; ric-4 was higher than that in either unc-64 or ric-4 single mutant nematodes, suggesting that unc-64 functions in parallel with ric-4 in regulating fat storage. CONCLUSION: The plasma membrane-associated SNAREs syntaxin/unc-64 and SNAP-25/ric-4 function in parallel in regulating fat storage in C. elegans, probably through the ARC105/mdt-15- and SREBP/sbp-1-mediated signaling pathways.

Optimization of riboflavin production by recombinant Bacillus subtilis RH44 using statistical designs.

A sequential optimization strategy, based on statistical experimental designs, was used to enhance the production of riboflavin by recombinant Bacillus subtilis RH44. In the first instance, the medium components were optimized in shake flask cultures. After preliminary experiments of nitrogen source selection, the two-level Plackett-Burman (PB) design was implemented to screen medium components that significantly influence riboflavin production. Among the 15 variables tested, glucose, NaNO(3), K(2)HPO(4), ZnSO(4), and MnCl(2) were identified as the most significant factors (confidence levels above 95%) for riboflavin production. The optimal values of these five variables were determined by response surface methodology (RSM) based on the central composite design (CCD). The validity of the model developed was verified, and the optimum medium led to a maximum riboflavin concentration of 6.65 g/l, which was 44.3 and 76.4% higher than the improved medium and the basal medium, respectively. A glucose-limited fed-batch culture profile in a 5-l fermentor was consequently designed according to the above optimum medium in shake flasks. A final riboflavin concentration of 16.36 g/l was obtained in 48 h, which further verified the practicability of this optimum strategy.