Effects of 24 Weeks of a Supervised Walk Training on Knee Muscle Strength and Quality of Life in Older Female Total Knee Arthroplasty: A Retrospective Cohort Study.

Poor supervision, impaired exercise adherence, and low compliance with exercise regimens result in inconsistent effects regarding exercise interventions. A supervised-walk training regimen (9 km/week) may have a positive effect on functional recovery in female total knee arthroplasty (TKA). This study aimed to evaluate the effect of a supervised walking regimen on lower limb muscle strength, functional fitness, and patient-reported outcomes in female TKA. Twenty-eight female TKA were allocated into a control (CON) (n = 14) or walk training (WT) (n = 14) group. WT on treadmills was initiated 12 weeks after TKA. All patients were examined for lower muscle strength (including extension and flexion of hip and knee), physical function (including a 6-min walk test, 8-foot up-and-go test, and 30-s chair stand test), and Knee Injury and Osteoarthritis Outcome Score (KOOS) questionnaire. Knee flexor (WT: CON; 64.4 ± 4.1 nm/kg: 43.7±3.3 nm/kg; p = 0.001; effect size: 5.62) and extensor strengths (WT: CON; 73.1 ± 7.5 nm/kg: 48.2 ± 2.4 nm/kg; p = 0.001; effect size: 4.47) statistically increased in the WT group compared to the CON group. The 6-min walk test (from 341.3 ± 20.5 m to 405.5 ± 30.7 m; p = 0.001; effect size: 2.46) and 8-foot up-and-go test (from 9.5 ± 0.7 s to 8.3 ± 0.7 s; p = 0.002; effect size: 1.71) tests also showed significant improvements in the WT group in the follow-up compared to the baseline. An increase in quality of life score according to the KOOS questionnaire (WT: CON; 91.0 ± 2.8: 68.1 ± 5.8; p = 0.001; effect size: 5.02) was noted in the WT group compared to the CON group in the follow-up. WT facilitated improvements in knee muscle strength and functional outcomes in TKA patients.

Inhibition of Neddylation Suppresses Osteoclast Differentiation and Function In Vitro and Alleviates Osteoporosis In Vivo.

Neddylation, or the covalent addition of NEDD8 to specific lysine residue of proteins, is a reversible posttranslational modification, which regulates numerous biological functions; however, its involvement and therapeutic significance in osteoporosis remains unknown. Our results revealed that during the soluble receptor activator of nuclear factor-κB ligand (sRANKL)-stimulated osteoclast differentiation, the neddylation and expression of UBA3, the NEDD8-activating enzyme (NAE) catalytic subunit, were dose- and time-dependently upregulated in RAW 264.7 macrophages. UBA3 knockdown for diminishing NAE activity or administering low doses of the NAE inhibitor MLN4924 significantly suppressed sRANKL-stimulated osteoclast differentiation and bone-resorbing activity in the macrophages by inhibiting sRANKL-stimulated neddylation and tumor necrosis factor receptor-associated factor 6 (TRAF6)-activated transforming growth factor-ß-activated kinase 1 (TAK1) downstream signaling for diminishing nuclear factor-activated T cells c1 (NFATc1) expression. sRANKL enhanced the interaction of TRAF6 with the neddylated proteins and the polyubiquitination of TRAF6's lysine 63, which activated TAK1 downstream signaling; however, this process was inhibited by MLN4924. MLN4924 significantly reduced osteoporosis in an ovariectomy- and sRANKL-induced osteoporosis mouse model in vivo. Our novel finding was that NAE-mediated neddylation participates in RANKL-activated TRAF6-TAK1-NFATc1 signaling during osteoclast differentiation and osteoporosis, suggesting that neddylation may be a new target for treating osteoporosis.

Characterization of Methicillin-Resistant Staphylococcus aureus Isolates from Periprosthetic Joint Infections.

Periprosthetic joint infection (PJI) is a troublesome clinical issue in total joint arthroplasty (TJA). Although methicillin-resistant Staphylococcus aureus (MRSA) is considered to be the most serious pathogen in PJIs, little is known about the genotypic and phenotypic characteristics of MRSA clones isolated from PJI patients. A total of 36 MRSA isolates from PJI patients were collected at the Chang-Gung Memorial Hospital in Taiwan from May 2016 to October 2019. All MRSA isolates were subjected to genome typing. The prevalence of Panton-Valentine leucocidin (PVL), the antibiotic susceptibility profile, and the biofilm formation ability were compared among different MRSA genogroups. Additionally, demographics and clinical manifestations of patients infected with different MRSA genogroups were investigated. Eight sequence types (STs) were identified among 36 isolated from PJIs. According to the incidence of MRSA genotypes in PJIs, in this study, we divided them into four groups, including ST8 (n = 10), ST59 (n = 8), ST239 (n = 11), and other STs (n = 7). For the antibiotic susceptibility testing, we found that all MRSA isolates in the ST239 group were highly resistant to ciprofloxacin, gentamicin trimethoprim-sulfamethoxazole, and levofloxacin. Additionally, ST239 MRSA also had a higher ability to form biofilm than other groups. Importantly, patients with ST239 infection typically had a fever and exhibited higher levels of inflammatory markers, including C-reactive protein (CRP) and white blood cell count (WBC). Epidemiological investigations revealed that knee PJIs were mainly attributed to infection with ST59 MRSA and increasing trends for infection with ST8 and other ST types of MRSAs in PJI patients were observed from 2016 to 2019. The identification of MRSA genotypes in PJIs may be helpful for the management of PJIs.

In Vitro and In Vivo Comparison of Bone Growth Characteristics in Additive-Manufactured Porous Titanium, Nonporous Titanium, and Porous Tantalum Interbody Cages.

Autogenous bone grafts are the gold standard for interbody fusion implant materials; however, they have several disadvantages. Tantalum (Ta) and titanium (Ti) are ideal materials for interbody cages because of their biocompatibility, particularly when they are incorporated into a three-dimensional (3D) porous structure. We conducted an in vitro investigation of the cell attachment and osteogenic markers of self-fabricated uniform porous Ti (20%, 40%, 60%, and 80%), nonporous Ti, and porous Ta cages (n = 6) in each group. Cell attachment, osteogenic markers, and alkaline phosphatase (ALP) were measured. An in vivo study was performed using a pig-posterior-instrumented anterior interbody fusion model to compare the porous Ti (60%), nonporous Ti, and porous Ta interbody cages in 12 pigs. Implant migration and subsidence, determined using plain radiographs, were recorded before surgery, immediately after surgery, and at 1, 3, and 6 months after surgery. Harvested implants were assessed for bone ingrowth and attachment. Relative to the 20% and 40% porous Ti cages, the 60% and 80% cages achieved superior cellular migration into cage pores. Among the cages, osteogenic marker and ALP activity levels were the highest in the 60% porous Ti cage, osteocalcin expression was the highest in the nonporous Ti cage, and the 60% porous Ti cage exhibited the lowest subsidence. In conclusion, the designed porous Ti cage is biocompatible and suitable for lumbar interbody fusion surgery and exhibits faster fusion with less subsidence compared with porous Ta and nonporous Ti cages.

Effect of resistance training on satellite cells in old mice - a transcriptome study : implications for sarcopenia.

AIMS: The decrease in the number of satellite cells (SCs), contributing to myofibre formation and reconstitution, and their proliferative capacity, leads to muscle loss, a condition known as sarcopenia. Resistance training can prevent muscle loss; however, the underlying mechanisms of resistance training effects on SCs are not well understood. We therefore conducted a comprehensive transcriptome analysis of SCs in a mouse model. METHODS: We compared the differentially expressed genes of SCs in young mice (eight weeks old), middle-aged (48-week-old) mice with resistance training intervention (MID+ T), and mice without exercise (MID) using next-generation sequencing and bioinformatics. RESULTS: After the bioinformatic analysis, the PI3K-Akt signalling pathway and the regulation of actin cytoskeleton in particular were highlighted among the top ten pathways with the most differentially expressed genes involved in the young/MID and MID+ T/MID groups. The expression of Gng5, Atf2, and Rtor in the PI3K-Akt signalling pathway was higher in the young and MID+ T groups compared with the MID group. Similarly, Limk1, Arhgef12, and Araf in the regulation of the actin cytoskeleton pathway had a similar bias. Moreover, the protein expression profiles of Atf2, Rptor, and Ccnd3 in each group were paralleled with the results of NGS. CONCLUSION: Our results revealed that age-induced muscle loss might result from age-influenced genes that contribute to muscle development in SCs. After resistance training, age-impaired genes were reactivated, and age-induced genes were depressed. The change fold in these genes in the young/MID mice resembled those in the MID + T/MID group, suggesting that resistance training can rejuvenate the self-renewing ability of SCs by recovering age-influenced genes to prevent sarcopenia. Cite this article: Bone Joint Res 2022;11(2):121-133.

Predicting osteoporosis with body compositions in postmenopausal women: a non-invasive method.

BACKGROUND: The prevalence of osteoporosis is rising steadily as the aging population increases. Bone mineral density (BMD) assessment is a golden standard to establish the diagnosis of osteoporosis. However, the accessibility and radiation exposure limited its role in community screening. A more convenient approach for screening is suggested. METHODS: A total of 363 postmenopausal women over the age of 50 were included in this study and assessed with the body composition [including fat-free mass (FFM), fat mass (FM), and basal metabolic rate (BMR)] and BMD. Normal distributions and correlation coefficients among variables were calculated using the Shapiro-Wilk test and Pearson's correlation analysis, respectively. A receiver operating characteristic (ROC) curve was plotted, and the area under ROC curves (AUC) was determined to obtain the optimal cutoff values of the body composition variables for osteoporosis prediction. RESULTS: The correlation coefficient of FFM, FM, FM ratio, and BMR with femur neck T-score was 0.373, 0.266, 0.165, and 0.369, respectively, while with spine T-score was 0.350, 0.251, 0.166, and 0.352, respectively (p < 0.01 for all). FFM, FM, and BMR showed an optimal cutoff value of 37.9 kg, 18.6 kg, and 1187.5 kcal, respectively, for detecting osteoporosis. CONCLUSIONS: The present study provided a model to predict osteoporosis in postmenopausal women, and the optimal cutoff value of FFM, FM, and BMR could be calculated in the Asian population. Among these factors, BMR seemed a better predictor than others. The BMR could be a target for exercise intervention in postmenopausal women for maintaining or improving BMD. TRIAL REGISTRATION: ClinicalTrials.gov , NCT02936336 . Retrospectively registered on13 October 2016.

Comparison of computer-assisted navigation and conventional instrumentation for bilateral total knee arthroplasty: The outcomes at mid-term follow-up.

It remains unclear if computer-assisted surgery (CAS) technique actually improves the clinical outcomes of total knee arthroplasty (TKA) and decreases the failure rate. The purpose of this retrospective study was to compare the functional results of TKA in a series of patients who underwent staged bilateral TKAs with CAS TKA in 1 knee and conventional TKA in the contralateral knee.From January 1997 to December 2010, we collected 60 patients who were randomly assigned to receive CAS TKA in 1 limb and conventional TKA in the other. The Brainlab Vector Vision navigation system was used for CAS TKA, and the DePuy press-fit condylar sigma guide system was used for conventional TKA. Patients were assessed before surgery, 3 months and 1 year after surgery, and annually thereafter. IKS criteria were used for radiographic evaluation. Clinical and functional evaluation using the scoring system of hospital for special surgery (HSS), international knee society (IKS), Western Ontario and McMaster University osteoarthritis index (WOMAC), and short form-36 (SF-36) were obtained on each knee, before surgery, and at each follow-up visit. Pertinent statistical methods were adopted for data analysis.Fifty-six patients were available for analysis and 44 of the patients were female. The mean duration of follow-up was 8.1 years. Less blood loss (P = .007) and longer operation time were noted for CAS TKAs when compared with conventional TKAs. Precise alignment and fewer outliers of the lower limb and prosthetic component positions were found for CAS TKAs (P < .001). There were no differences between the 2 groups before surgery and at the latest follow-up with regard to scores for HSS, IKS, WOMAC, and SF-36 as well as active range of motion.The clinical outcomes of CAS TKAs at the 8-year follow-up were similar to those of conventional TKAs despite the better radiographic alignment and fewer outliers achieved with navigation assistance.

Twenty-four-week hospital-based progressive resistance training on functional recovery in female patients post total knee arthroplasty.

BACKGROUND: After total knee arthroplasty (TKA) surgery, a decline in muscle strength is associated with a decrease in function. The aim of this study was to demonstrate the effect of a further 24 weeks of hospital-based resistance training under supervision, and precise dose on knee functional recovery and daily activities for female TKA patients. METHODS: Twenty-nine patients who underwent unilateral primary TKA were allocated into either resistance training (RT) (n = 14) or control (CON) (n = 15) groups. All patients were assessed, with an isokinetic dynamometer, for hip and knee flexor and extensor muscle strength, physical function test, and Knee Injury and Osteoarthritis Outcome Score (KOOS). Resistance training was initiated three months after index surgery. The assessments were performed before exercise (Baseline), in the middle of the resistance training at 12 weeks (Mid-exercise), completion of the resistance training (Post-exercise), and 12 weeks after resistance training completion (Follow-up). A statistical test was performed by using generalized estimating equations. RESULTS: Patients in RT had more of an increase in both knee extensor and flexor muscle strength than those in CON at the Post-exercise assessment. The six-minute walk test distance was more in RT compared with CON at the same Post-exercise assessment. Furthermore, the RT group had increases in Activities of daily living and Sports subscales compared to the CON group. CONCLUSIONS: A further 24 weeks of hospital-based progressive resistance training facilitated improvement in knee muscle strength and functional outcome in TKA patients. Active hospital-based progressive resistance training is recommended for rehabilitation following TKA surgery.

The effects of high impact exercise intervention on bone mineral density, physical fitness, and quality of life in postmenopausal women with osteopenia: A retrospective cohort study.

Osteoporosis and osteopenia prevailed in postmenopausal women and predisposed to osteoporotic fractures that increase mortality, morbidity, and the cost of social care. Here, we investigated the effect of 24 weeks of aerobic dancing on the bone miner density, physical fitness and health-related quality of life (HRQoL) in postmenopausal women with osteopenia. Total 80 participants (control [CON]: 40; exercise [EX]: 40) were included in the final analysis. The EX group underwent a 24-week aerobic dance intervention. Bone mineral density (BMD), physical fitness, and SF-36 questionnaire were assessed at baseline and 24-weeks. The BMD change in the femoral neck at the 24-weeks were significantly different between the 2 groups (CON: -1.3 ±â€Š2.7%, EX: 3.1 ±â€Š4.6%, P = .001). Grip strength, sidestep and physical functional domain of HRQoL in the EX group were significantly improved compared to the CON. The results were suggested 24-week aerobic dance intervention could result in the lower the incidence of bone fracture through increasing BMD and decreasing fall risk for postmenopausal women.

Correction: Evaluation of a novel biodegradable thermosensitive keto-hydrogel for improving postoperative pain in a rat model.

[This corrects the article DOI: 10.1371/journal.pone.0186784.].

Evaluation of a novel biodegradable thermosensitive keto-hydrogel for improving postoperative pain in a rat model.

This study evaluates the sustained analgesic effect of ketorolac-eluting thermosensitive biodegradable hydrogel in the plantar incisional pain model of the rat hind-paw. A ketorolac-embedded 2, 2'-Bis (2-oxazolin) (BOX) linking methoxy-poly(ethylene glycol) and poly(lactide-co-glycolide) (mPEG-PLGA) diblock copolymer (BOX copolymer) was synthesized as keto-hydrogel based on optimal sol-gel phase transition and in vitro drug release profile. The effect of keto-hydrogel on postoperative pain (POP) was assessed using the established plantar incisional pain model in hind-paw of rats and compared to that of ketorolac solution. Pain and sensory threshold, as well as pain scoring, were evaluated with behavioral tests by means of anesthesiometer and incapacitance apparatus, respectively. Pro-inflammatory cytokine levels (TNF-α, IL-6, VEGF, and IL-1ß) around incisional wounds were measured by ELISA. Tissue histology was assessed using hematoxylin and eosin and Masson's trichrome staining. Ten mg/mL (25 wt%) keto-hydrogel showed a sol-gel transition at 26.4°C with a 10-day sustained drug release profile in vitro. Compared to ketorolac solution group, the concentration of ketorolac in tissue fluid was higher in the keto-hydrogel group during the first 18 h of application. Keto-hydrogel elevated pain and sensory threshold, increased weight-bearing capacity, and significantly reduced the levels of TNF-α, IL-6, and IL-1ß while enhanced VEGF in tissue fluid. Histologic analysis reveals greater epithelialization and collagen deposition around wound treated with keto-hydrogel. In conclusion, our study suggests that keto-hydrogel is an ideal compound to treat POP with a secondary gain of improved incisional wound healing.

Circuit training enhances function in patients undergoing total knee arthroplasty: a retrospective cohort study.

BACKGROUND: The number of patients receiving total knee arthroplasty (TKA) has been rising every year due to the aging population and the obesity epidemic. Post-operative rehabilitation is important for the outcome of TKA. METHODS: A series of 34 patients who underwent primary unilateral TKA was retrospectively collected and divided into either exercise group (n = 16) and control group (n = 18). The exercise group underwent a 24-week course of circuit training beginning 3 months after total knee arthroplasty (TKA). The effect of circuit training on TKA patients in terms of motion analysis, muscle strength testing, Knee injury and Osteoarthritis Outcomes Score (KOOS) questionnaire and patient-reported outcome measurement Short-Form Health Survey (SF-36) at the pre-operation, pre-exercise, mid-exercise, and post-exercise. RESULTS: Motion analysis revealed the stride length, step velocity, and excursion of active knee range of motion significantly improved in the exercise group when compared to those in the control group. KOOS questionnaire showed a greater improvement in pain, ADL, and total scores in the exercise group. The SF-36 questionnaire revealed a significant improvement in general health, bodily pain, social function, and physical components score in the exercise group. CONCLUSIONS: The post-operative circuit training intervention can facilitate recovery of knee function and decrease the degree of pain in the TKA and might be considered a useful adjunct rehabilitative modality. The ultimate influence of circuit training on TKA needs further a prospective randomized clinical trial study and long-term investigation. TRIAL REGISTRATION: NCT02928562.

The Chinese Herbal Mixture Tien-Hsien Liquid Augments the Anticancer Immunity in Tumor Cell-Vaccinated Mice.

BACKGROUND: The Chinese herbal mixture, Tien-Hsien liquid (THL), has been used as an anticancer dietary supplement for more than 20 years. Our previous studies have shown that THL can modulate immune responseand inhibit tumor growth. In this study, we further evaluated the effect of THL on anticancer immune response in mice vaccinated with γ-ray-irradiated tumor cells. METHODS: The antitumor effect of THL was determined in mice vaccinated with low-tumorigenic CT-26-low colon cancer cells or γ-ray-irradiated high-tumorigenic CT-26-high colon cancer cells. The number of natural killer (NK) cells and T lymphocytes in the spleen was analyzed by flow cytometry. The tumor-killing activities of NK cells and cytotoxic T lymphocytes (CTLs) were analyzed by flow cytometry using YAC-1 and CT-26-high cells, respectively, as target cells. The levels of IFN-γ, IL-2, and TNF-α were determined by ELISA. RESULTS: THL suppressed the growth of CT-26-high tumor in mice previously vaccinated with low-tumorigenic CT-26-low cells or γ-irradiated CT-26-high cells. THL increased the populations of NK cells and CD4+ T lymphocytes in the spleen and enhanced the tumor-killing activities of NK cells and CTL in mice vaccinated with γ-irradiated CT-26-high cells. THL increased the production of IFN-γ, IL-2, and TNF-α in mice vaccinated with γ-irradiated CT-26-high cells. CONCLUSION: THL can enhance the antitumor immune responses in mice vaccinated with killed tumor cells. These results suggest that THL may be used as a complementary medicine for cancer patients previously treated with killed tumor cell vaccines, radiotherapy, or chemotherapy.

Network Signatures of IgG Immune Repertoires in Hepatitis B Associated Chronic Infection and Vaccination Responses.

The repertoire of IgG antibody responses to infection and vaccination varies depending on the characteristics of the immunogen and the ability of the host to mount a protective immune response. Chronic hepatitis B virus (HBV) infections are marked by persistent infection and immune tolerance to vaccination. This disease offers a unique opportunity to discover key repertoire signatures during infection and in response to vaccination. Complementarity determining region 3 of an antibody heavy chain (CDR-H3) has a major impact on the antigenic specificity of an antibody. We used next-generation sequencing to characterize the CDR-H3 sequences in paired siblings of 4 families in which only one member of each pair had chronic HBV infection. Blood samples were obtained before and 2 weeks after HBV vaccination. The analysis revealed a huge network of sequence-related CDR-H3 clones found almost exclusively among carriers. In contrast, vaccination induced significant increases of CDR-H3 cluster diversities among siblings without hepatitis B. Several vaccination-associated clone clusters were identified. Similar findings of vaccination-associated clone networks were observed in healthy adults receiving HBV boosters. These strategies can be used to identify signatures of other infectious diseases and accelerate discoveries of antibody sequences with important biomedical implications.

Cellular protein HAX1 interacts with the influenza A virus PA polymerase subunit and impedes its nuclear translocation.

Transcription and replication of the influenza A virus RNA genome occur in the nucleus through the viral RNA-dependent RNA polymerase consisting of PB1, PB2, and PA. Cellular factors that associate with the viral polymerase complex play important roles in these processes. To look for cellular factors that could associate with influenza A virus PA protein, we have carried out a yeast two-hybrid screen using a HeLa cell cDNA library. We identified six cellular proteins that may interact with PA. We focused our study on one of the new PA-interacting proteins, HAX1, a protein with antiapoptotic function. By using glutathione S-transferase pulldown and coimmunoprecipitation assays, we demonstrate that HAX1 specifically interacts with PA in vitro and in vivo and that HAX1 interacts with the nuclear localization signal domain of PA. Nuclear accumulation of PA was increased in HAX1-knockdown cells, and this phenotype could be reversed by reexpression of HAX1, indicating that HAX1 can impede nuclear transport of PA. As a consequence, knockdown of HAX1 resulted in a significant increase in virus yield and polymerase activity in a minigenome assay, and this phenotype could be reversed by reexpression of HAX1, indicating that HAX1 can inhibit influenza A virus propagation. Together, these results not only provide insight into the mechanism underlying nuclear transport of PA but also identify an intrinsic host factor that restricts influenza A virus infection.

Fermentation product of soybean, black bean, and green bean mixture induces apoptosis in a wide variety of cancer cells.

Previous studies have shown that soybean fermentation products can act as cancer chemoprevention or therapeutic agents. In this study, the anticancer activities of a fermentation product of soybean, black bean, and green bean mixture (BN999) were investigated. We found that BN999 inhibited the growth of human breast cancer AU565 cells and prostate adenocarcinoma PC-3 cells but not that of normal human cells. BN999 induced apoptosis in various human cancer cells but not in normal human cells. BN999 treatment of AU565 cancer cells resulted in activation of calpain and caspase-8, -9, and -3, suggesting that BN999 induces apoptosis via receptor-, mitochondria-, and endoplasmic reticulum-mediated pathways. Finally, we showed that BN999 inhibited the growth of mouse CT-26 colon cancer xenografts in syngenic BALB/c mice without causing obvious side effects. Together, these data suggest that BN999 has potential to be used as a cancer chemoprevention or therapeutic agent.

Inhibition of metastasis, angiogenesis, and tumor growth by Chinese herbal cocktail Tien-Hsien Liquid.

BACKGROUND: Advanced cancer is a multifactorial disease that demands treatments targeting multiple cellular pathways. Chinese herbal cocktail which contains various phytochemicals may target multiple dys-regulated pathways in cancer cells and thus may provide an alternative/complementary way to treat cancers. Previously we reported that the Chinese herbal cocktail Tien-Hsien Liguid (THL) can specifically induce apoptosis in various cancer cells and have immuno-modulating activity. In this study, we further evaluated the anti-metastatic, anti-angiogenic and anti-tumor activities of THL with a series of in vitro and in vivo experiments. METHODS: The migration and invasion of cancer cells and endothelial cells was determined by Boyden chamber transwell assays. The effect of THL on pulmonary metastasis was done by injecting CT-26 colon cancer cells intravenously to syngenic mice. The in vitro and in vivo microvessel formation was determined by the tube formation assay and the Matrigel plug assay, respectively. The in vivo anti-tumor effect of THL was determined by a human MDA-MB-231 breast cancer xenograft model. The expression of metalloproteinase (MMP)-2, MMP-9, and urokinase plasminogen activator (uPA) was measured by gelatin zymography. The expression of HIF-1alpha and the phosphorylation of ERK1/2 were determined by Western blot. RESULTS: THL inhibited the migration and invasion ability of various cancer cells in vitro, decreased the secretion of MMP-2, MMP-9, and uPA and the activity of ERK1/2 in cancer cells, and suppressed pulmonary metastasis of CT-26 cancer cells in syngenic mice. Moreover, THL inhibited the migration, invasion, and tube formation of endothelial cells in vitro, decreased the secretion of MMP-2 and uPA in endothelial cells, and suppressed neovascularization in Matrigel plugs in mice. Besides its inhibitory effect on endothelial cells, THL inhibited hypoxia-induced HIF-1alpha and vascular endothelial growth factor-A expression in cancer cells. Finally, our results show that THL inhibited the growth of human MDA-MB-231 breast cancer xenografts in NOD-SCID mice. This suppression of tumor growth was associated with decreased microvessel formation and increased apoptosis caused by THL. CONCLUSION: Our data demonstrate that THL had broad-spectra anti-cancer activities and merits further evaluation for its use in cancer therapy.

Epstein-Barr virus nuclear antigen 2 disrupts mitotic checkpoint and causes chromosomal instability.

The Epstein-Barr virus nuclear antigen 2 (EBNA2) plays a key role in transformation of B-lymphocytes mediated by Epstein-Barr virus (EBV) and can induce tumor formation in transgenic mice. However, the precise mechanism underlying EBNA2-mediated tumorigenesis remains elusive. Here, we report that EBNA2 can compromise mitotic spindle checkpoint (MSC) induced by the spindle inhibitor nocodazole and cause chromosomal instability (CIN) in HEp-2, U2-OS and BJAB cells. When EBNA2-expressing cells were treated with nocodazole, they exited mitosis prematurely and initiated another round of DNA synthesis. Nucleolocalization of EBNA2 was essential for EBNA2 to compromise MSC and to cause CIN. The metaphase chromosome spread data indicated that the EBNA2-expressing U2-OS cells showed a more heterogenous chromosome number distribution than the vector-transfected and parental cells. The median chromosome number for EBNA2-expressing, vector-transfected and parental U2-OS cells is 75, 65 and 64, respectively. EBNA2 was shown to be able to downregulate mitotic arrest deficient 2 (MAD2) approximately 2- to 3-fold and upregulate polo-like kinase 1 (PLK1) approximately 2-fold. The dysregulation of MAD2 and PLK1 may lead to activation of anaphase promoting complex/cyclosome and premature degradation of securin. Indeed, we found that when MSC was induced by nocodazole, securin was prematurely degraded in EBNA2-expressing cells. Finally, we show that EBNA2 could induce micronuclei and multinuclei formation in HEp-2 and U2-OS cells. Together, these studies reveal a new function of EBNA2 in cell-cycle regulation and may shed light on the role of EBNA2 in EBV-mediated tumorigenesis.

Role of RppA in the regulation of polymyxin b susceptibility, swarming, and virulence factor expression in Proteus mirabilis.

Proteus mirabilis, a human pathogen that frequently causes urinary tract infections, is intrinsically highly resistant to cationic antimicrobial peptides, such as polymyxin B (PB). To explore the mechanisms underlying P. mirabilis resistance to PB, a mutant which displayed increased (> 160-fold) sensitivity to PB was identified by transposon mutagenesis. This mutant was found to have Tn5 inserted into a novel gene, rppA. Sequence analysis indicated that rppA may encode a response regulator of the two-component system and is located upstream of the rppB gene, which may encode a membrane sensor kinase. An rppA knockout mutant of P. mirabilis had an altered lipopolysaccharide (LPS) profile. The LPS purified from the rppA knockout mutant could bind more PB than the LPS purified from the wild type. These properties of the rppA knockout mutant may contribute to its PB-sensitive phenotype. The rppA knockout mutant exhibited greater swarming motility and cytotoxic activity and expressed higher levels of flagellin and hemolysin than the wild type, suggesting that RppA negatively regulates swarming, hemolysin expression, and cytotoxic activity in P. mirabilis. PB could modulate LPS synthesis and modification, swarming, hemolysin expression, and cytotoxic activity in P. mirabilis through an RppA-dependent pathway, suggesting that PB could serve as a signal to regulate RppA activity. Finally, we demonstrated that the expression of rppA was up-regulated by a low concentration of PB and down-regulated by a high concentration of Mg2+. Together, these data highlight the essential role of RppA in regulating PB susceptibility and virulence functions in P. mirabilis.