Acupuncture in Sports Medicine.

Acupuncture is gaining popularity and wider acceptance as a treatment modality within the field of sports medicine. Our objective was to provide a comprehensive review of the existing literature pertaining to acupuncture in sports medicine to shed light on approaches utilized in acupuncture while revealing its personalized nature and its impact on athletes' preparation, performance, and recovery. We evaluated acupuncture research in the context of medicine and sports-related injury treatment, assessing its impact on athletic performance across demographics of athletes. Athletes participating in most sports have shown positive outcomes from acupuncture interventions. Acupuncture improves peak oxygen levels, maximum heart rate, delayed-onset muscle soreness, pain, swelling, explosive force production, and joint mobility. Furthermore, the efficacy of acupuncture appears to be similar regardless of age and sex. Lastly, the acceptance of acupuncture is influenced by cultural factors, with Western and traditional East Asian cultures exhibiting distinct perspectives on its rationale and mechanisms of action. Traditional East Asian acupuncturists typically employ qi and meridian theories in their acupuncture practices, with the recent incorporation of Western concepts. Acupuncture shows promise as an effective treatment for musculoskeletal pain and neuropathies in athletes across different age groups and for addressing injuries in various sports. Our comprehensive review will enhance our understanding of acupuncture's potential as a complementary or distinct therapeutic approach compared to conventional therapies. Additionally, our review explores its specific applications within different sports and delves into the cultural dimensions involved in integrating this practice into modern sports medicine.

Do In-Hospital Rothman Index Scores Predict Postdischarge Adverse Events and Discharge Location After Total Knee Arthroplasty?

BACKGROUND: There have been efforts to reduce adverse events and unplanned readmissions after total joint arthroplasty. The Rothman Index (RI) is a real-time, composite measure of medical acuity for hospitalized patients. We aimed to examine the association among in-hospital RI scores and complications, readmissions, and discharge location after total knee arthroplasty (TKA). We hypothesized that RI scores could be used to predict the outcomes of interest. METHODS: This is a retrospective study of an institutional database of elective, primary TKA from July 2018 until December 2019. Complications and readmissions were defined per Centers for Medicare and Medicaid Services. Analysis included multivariate regression, computation of the area under the curve (AUC), and the Youden Index to set RI thresholds. RESULTS: The study cohort's (n = 957) complications (2.4%), readmissions (3.6%), and nonhome discharge (13.7%) were reported. All RI metrics (minimum, maximum, last, mean, range, 25th%, and 75th%) were significantly associated with increased odds of readmission and home discharge (all P < .05). RI scores were not significantly associated with complications. The optimal RI thresholds for increased risk of readmission were last ≤ 71 (AUC = 0.65), mean ≤ 67 (AUC = 0.66), or maximum ≤ 80 (AUC = 0.63). The optimal RI thresholds for increased risk of home discharge were minimum ≥ 53 (AUC = 0.65), mean ≥ 69 (AUC = 0.65), or maximum ≥ 81 (AUC = 0.60). CONCLUSION: RI values may be used to predict readmission or home discharge after TKA.

An inhibitory hippocampal-thalamic pathway modulates remote memory retrieval.

Memories are supported by distributed hippocampal-thalamic-cortical networks, but the brain regions that contribute to network activity may vary with memory age. This process of reorganization is referred to as systems consolidation, and previous studies have examined the relationship between the activation of different hippocampal, thalamic, and cortical brain regions and memory age at the time of recall. While the activation of some brain regions increases with memory age, other regions become less active. In mice, here we show that the active disengagement of one such brain region, the anterodorsal thalamic nucleus, is necessary for recall at remote time-points and, in addition, which projection(s) mediate such inhibition. Specifically, we identified a sparse inhibitory projection from CA3 to the anterodorsal thalamic nucleus that becomes more active during systems consolidation, such that it is necessary for contextual fear memory retrieval at remote, but not recent, time-points post-learning.

Minimally disruptive medicine (MDM) in clinical practice: a qualitative case study of the human immunodeficiency virus (HIV) clinic care model.

BACKGROUND: Recent evidence suggests the need to reframe healthcare delivery for patients with chronic conditions, with emphasis on minimizing healthcare footprint/workload on patients, caregivers, clinicians and health systems through the proposed Minimally Disruptive Medicine (MDM) care model named. HIV care models have evolved to further focus on understanding barriers and facilitators to care delivery while improving patient-centered outcomes (e.g., disease progression, adherence, access, quality of life). It is hypothesized that these models may provide an example of MDM care model in clinic practice. Therefore, this study aimed to observe and ascertain MDM-concordant and discordant elements that may exist within a tertiary-setting HIV clinic care model for patients living with HIV or AIDS (PLWHA). We also aimed to identify lessons learned from this setting to inform improving the feasibility and usefulness of MDM care model. METHODS: This qualitative case study occurred in multidisciplinary HIV comprehensive-care clinic within an urban tertiary-medical center. Participants included Adult PLWHA and informal caregivers (e.g. family/friends) attending the clinic for regular appointments were recruited. All clinic staff were eligible for recruitment. Measurements included; semi-guided interviews with patients, caregivers, or both; semi-guided interviews with varied clinicians (individually); and direct observations of clinical encounters (patient-clinicians), as well as staff daily operations in 2015-2017. The qualitative-data synthesis used iterative, mainly inductive thematic coding. RESULTS: Researcher interviews and observations data included 28 patients, 5 caregivers, and 14 care-team members. With few exceptions, the clinic care model elements aligned closely to the MDM model of care through supporting patient capacity/abilities (with some patients receiving minimal social support and limited assistance with reframing their biography) and minimizing workload/demands (with some patients challenged by the clinic hours of operation). CONCLUSIONS: The studied HIV clinic incorporated many of the MDM tenants, contributing to its validation, and informing gaps in knowledge. While these findings may support the design and implementation of care that is both minimally disruptive and maximally supportive, the impact of MDM on patient-important outcomes and different care settings require further studying.

Health and Wellness Coaching in Serving the Needs of Today's Patients: A Primer for Healthcare Professionals.

The past six decades have been marked by leaps and bounds in medical advances, while concurrently clinical outcomes and the quality of life continued to lag or decline. There is a need for more comprehensive approaches to delivering healthcare to patients that address illness and wellness within and outside healthcare settings. Mounting evidence shows that making sustainable changes in healthcare requires approaching patients'/individuals' care as a continuum-within and outside healthcare settings-while addressing their capacity (ie ability) and workload (ie demands) and incorporating their values and preferences. Health and Wellness Coaching (HWC) has been proposed as a solution to create partnerships to empower individuals to take ownership, leadership, and accountability of their well-being, using nondirective, empathic, and mindful conversations that employ motivational-interviewing and evidence-based approaches. Insufficient clarity exists among healthcare professionals in understanding the definition, roles, and types of HWC. This primer summarizes HWC concepts and history and compares HWC types and its potential role in promoting, supporting, and improving the well-being, clinical outcomes, and quality of life of the pertinent stakeholders. This primer also highlights current and potential areas of application of HWC within different subpopulations and healthcare-related settings.

Current trends in health coaching for chronic conditions: A systematic review and meta-analysis of randomized controlled trials.

BACKGROUND: Chronic conditions are placing a serious burden on individuals as well as the health care system. Health coaching (HC) has emerged as a promising approach that can support effective lifestyle interventions for chronic conditions. However, until now there is no particularly comprehensive systematic review of HC impact on a chronic condition from the angle of patient improvement and detail coaching characteristics reported. OBJECTIVE: To synthesize available studies on the efficacy and current status of HC interventions on the health of chronically ill adult patients. METHODS: The literature search will be conducted for trials published in English within the past four years. Electronic databases CINAHL, Cochrane Library, Embase, MEDLINE, and Scopus will be searched with keywords describing HC for chronic diseases. Randomized controlled trials that compare HC interventions to conventional care or other alternative therapies will be included. Data extraction will be conducted by two reviewers independently, and enrolled trials will be evaluated for quality and bias assessment. If appropriate, meta-analysis will be conducted on the last stage of the review; otherwise, the study findings will be described narratively. The software Review Manager (Revman version 5.3.5.) provided by the Cochrane Collaboration will be applied for the meta-analysis. RESULTS: This is the first study to comprehensively explore the effectiveness and current status of HC intervention for patients with chronic conditions. DISCUSSION: Study findings from this review will advance the appropriate utilization of coaching practice by determining whether HC is effective and feasible among patients with chronic disease. If proven effective, this approach may be applied more broadly through public health interventions. The current status findings will also provide evidence to inform decisions for integrating HC interventions into the current management pathway for individuals with chronic conditions. SYSTEMATIC REVIEW REGISTRATION: PROSPERO CRD42020153280.

Pathogenesis, Clinical Signs and Treatment Recommendations in Brittle Nails: A Review.

Nail plate brittleness (or fragility) is a common complaint affecting up to 20% of the population, especially women over 50 years of age, with fingernail fragility being more prevalent than toenail fragility. Nail brittleness is characterized by nails that split, flake and crumble, become soft and lose elasticity. The main clinical presentations are: onychoschizia, onychorrhexis, superficial granulation of keratin and worn-down nails. According to causative factors, we can distinguish 2 forms of nail fragility (NF): a primary "idiopathic or brittle nail syndrome" form and NF secondary to different causes such as inflammatory nail disorders, infections, systemic diseases and general conditions, traumas and alteration of the nail hydration. Optimal management requires treatment of the primary cause of brittle nails, when possible. In idiopathic NF oral supplementation, vitamins (especially biotin, also known as vitamin B7), trace elements and amino acids (especially cysteine) have been reported to be useful. In addition, several products, such as topical moisturizers and lacquers could be considered to restructure the affected nail plate and to reduce psychological impacts of this common problem.

Project ESTEEM protocol: a randomized controlled trial of an LGBTQ-affirmative treatment for young adult sexual minority men's mental and sexual health.

BACKGROUND: Young gay and bisexual men disproportionately experience depression, anxiety, and substance use problems and are among the highest risk group for HIV infection in the U.S. Diverse methods locate the source of these health disparities in young gay and bisexual men's exposure to minority stress. In fact, minority stress, psychiatric morbidity, substance use, and HIV risk fuel each other, forming a synergistic threat to young gay and bisexual men's health. Yet no known intervention addresses minority stress to improve mental health, substance use problems, or their joint impact on HIV risk in this population. This paper describes the design of a study to test the efficacy of such an intervention, called ESTEEM (Effective Skills to Empower Effective Men), a 10-session skills-building intervention designed to reduce young gay and bisexual men's co-occurring health risks by addressing the underlying cognitive, affective, and behavioral pathways through which minority stress impairs health. METHODS: This study, funded by the National Institute of Mental Health, is a three-arm randomized controlled trial to examine (1) the efficacy of ESTEEM compared to community mental health treatment and HIV counseling and testing and (2) whether ESTEEM works through its hypothesized cognitive, affective, and behavioral minority stress processes. Our primary outcome, measured 8 months after baseline, is condomless anal sex in the absence of PrEP or known undetectable viral load of HIV+ primary partners. Secondary outcomes include depression, anxiety, substance use, sexual compulsivity, and PrEP uptake, also measured 8 months after baseline. DISCUSSION: Delivering specific stand-alone treatments for specific mental, behavioral, and sexual health problems represents the current state of evidence-based practice. However, dissemination and implementation of this one treatment-one problem approach has not been ideal. A single intervention that reduces young gay and bisexual men's depression, anxiety, substance use, and HIV risk by reducing the common minority stress pathways across these problems would represent an efficient, cost-effective alternative to currently isolated approaches, and holds great promise for reducing sexual orientation health disparities among young men. TRIAL REGISTRATION: Registered October 10, 2016 to ClinicalTrials.gov Identifier: NCT02929069 .

Aquatic Therapy for People with Lymphedema: A Systematic Review and Meta-analysis.

BACKGROUND: Aquatic therapy has several proposed benefits for people with lymphedema. A systematic review of the evidence for aquatic therapy in lymphedema management has not been conducted. METHOD AND RESULTS: Systematic review and meta-analysis were conducted. Five electronic databases were searched to identify randomized controlled trials (RCTs) of people with lymphedema, which compared aquatic therapy with other lymphedema interventions. Qualitative analysis was undertaken where quantitative analysis was not possible. Study quality was assessed using physiotherapy evidence database (PEDro) scores. The strength of evidence was evaluated using the Grades of Recommendations Assessment, Development and Evaluation (GRADE) approach. Four RCTs of moderate quality (average PEDro score 6.5/10) were included in the review. Two studies provided results for inclusion in meta-analysis. There was moderate-level evidence of no significant short-term differences in lymphedema status (as measured by lymphedema relative volume) between patients who completed aqua lymphatic therapy (ALT) compared to land-based standard care (standardized mean difference [SMD]: 0.14; 95% confidence interval [CI]: -0.37 to 0.64, I2 = 0%, p = 0.59); and low-quality evidence of no significant difference between ALT and standard care for improving upper limb (UL) physical function (SMD -0.27, 95% CI: -0.78 to 0.23, I2 = 0%, p = 0.29). No adverse events reported. CONCLUSIONS: Current evidence indicates no significant benefit of ALT over standard land-based care for improving lymphedema status or physical function in people with UL lymphedema. Patient preference should guide the choice of care to facilitate adherence. Further research is required to strengthen the evidence from four studies in people with UL lymphedema, and to establish the efficacy of this intervention in people with lower limb lymphedema. Review registration: PROSPERO (CRD42015019900).

Simulation-based Assessment of the Management of Critical Events by Board-certified Anesthesiologists.

BACKGROUND: We sought to determine whether mannequin-based simulation can reliably characterize how board-certified anesthesiologists manage simulated medical emergencies. Our primary focus was to identify gaps in performance and to establish psychometric properties of the assessment methods. METHODS: A total of 263 consenting board-certified anesthesiologists participating in existing simulation-based maintenance of certification courses at one of eight simulation centers were video recorded performing simulated emergency scenarios. Each participated in two 20-min, standardized, high-fidelity simulated medical crisis scenarios, once each as primary anesthesiologist and first responder. Via a Delphi technique, an independent panel of expert anesthesiologists identified critical performance elements for each scenario. Trained, blinded anesthesiologists rated video recordings using standardized rating tools. Measures included the percentage of critical performance elements observed and holistic (one to nine ordinal scale) ratings of participant's technical and nontechnical performance. Raters also judged whether the performance was at a level expected of a board-certified anesthesiologist. RESULTS: Rater reliability for most measures was good. In 284 simulated emergencies, participants were rated as successfully completing 81% (interquartile range, 75 to 90%) of the critical performance elements. The median rating of both technical and nontechnical holistic performance was five, distributed across the nine-point scale. Approximately one-quarter of participants received low holistic ratings (i.e., three or less). Higher-rated performances were associated with younger age but not with previous simulation experience or other individual characteristics. Calling for help was associated with better individual and team performance. CONCLUSIONS: Standardized simulation-based assessment identified performance gaps informing opportunities for improvement. If a substantial proportion of experienced anesthesiologists struggle with managing medical emergencies, continuing medical education activities should be reevaluated.

Massage Therapy and Quality of Life in Osteoarthritis of the Knee: A Qualitative Study.

Objective: We hypothesized that participants receiving Swedish massage would experience benefits such as stress reduction and enhanced quality of life, in addition to the osteoarthritis-specific effects assessed in a randomized controlled clinical trial. Design: Qualitative methods were used to explore a deeper contextual understanding of participants' experiences with massage and osteoarthritis, in addition to the quantitative data collected from primary and secondary outcome measures of the dose-finding study. Setting: Two community hospitals affiliated with academic health centers in Connecticut and New Jersey. Subjects: Eighteen adults who previously participated in a dose-finding clinical trial of massage therapy for osteoarthritis of the knee. Methods: Face-to-face and telephone interviews using a standardized interview guide. Triangulation of qualitative and quantitative data allowed for a more thorough understanding of the effects of massage therapy. Results: Three salient themes emerged from our analysis. Participants discussed 1) relaxation effects, 2) improved quality of life associated with receiving massage therapy, and 3) the accessibility of massage therapy in treating osteoarthritis. Conclusions: Participant responses noted empowerment with an improved ability to perform activities of daily living after experiencing massage therapy. The majority of statements were consistent with their quantitative changes on standard osteoarthritis measures. Future research in pain conditions should include health-related quality of life assessments as well as outcomes related to perceived well-being, along with greater exploration of the concept of salutogenic side effects of an intervention in the context of complementary and integrative therapies.

Pilot study of massage in veterans with knee osteoarthritis.

OBJECTIVES: To (1) assess the feasibility and acceptability of Swedish massage among Department of Veterans Affairs (VA) health care users with knee osteoarthritis (OA) and (2) collect preliminary data on efficacy of Swedish massage in this patient group. DESIGN: Experimental pilot study. SETTING: Duke Integrative Medicine clinic and VA Medical Center, Durham, North Carolina. PATIENTS: Twenty-five veterans with symptomatic knee OA. INTERVENTIONS: Eight weekly 1-hour sessions of full-body Swedish massage. OUTCOME MEASURES: Primary: Western Ontario and McMaster University Osteoarthritis Index (WOMAC) and global pain (Visual Analog Scale [VAS]). Secondary: National Institutes of Health Patient Reported Outcomes Measurement Information System-Pain Interference Questionnaire 6b (PROMIS-PI 6b), 12-Item Short-Form Health Survey (SF-12 v1) and the EuroQol health status index (EQ-5D-5L), knee range of motion (ROM), and time to walk 50 feet. RESULTS: Study feasibility was established by a 92% retention rate with 99% of massage visits and 100% of research visits completed. Results showed significant improvements in self-reported OA-related pain, stiffness and function (30% improvement in Global WOMAC scores; p=0.001) and knee pain over the past 7 days (36% improvement in VAS score; p<0.001). PROMIS-PI, EQ-5D-5L, and physical composite score of the SF-12 also significantly improved (p<0.01 for all), while the mental composite score of the SF-12 and knee ROM showed trends toward significant improvement. Time to walk 50 feet did not significantly improve. CONCLUSIONS: Results of this pilot study support the feasibility and acceptability of Swedish massage among VA health care users as well as preliminary data suggesting its efficacy for reducing pain due to knee OA. If results are confirmed in a larger randomized trial, massage could be an important component of regular care for these patients.

Orally administered novel cyclic pentapeptide P-317 alleviates symptoms of diarrhoea-predominant irritable bowel syndrome.

OBJECTIVE: The aim of our study was to characterize the effect of P-317, a novel cyclic derivative of morphiceptin, on gastrointestinal (GI) motility and abdominal pain in mouse models mimicking symptoms of diarrhoea-predominant irritable bowel syndrome (IBS-D). METHODS: The effect of P-317 on mouse intestinal motility was characterized in vitro and in vivo in physiological and pathophysiological conditions. The antinociceptive action of P-317 was characterized in the mustard oil-induced abdominal pain model and the writhing test. Locomotor activity and grip-strength tests were used to evaluate the effect of P-317 in the central nervous system (CNS). To translate our study to clinical conditions, the semi-quantitative expression of µ-opioid receptors (MOP) and κ-opioid receptors (KOP) messenger RNA (mRNA) in human colonic samples from IBS-D patients was quantified. KEY FINDINGS: In vitro, P-317 (10(-10) -10(-6) M) inhibited colonic and ileal smooth muscle contractions in a concentration-dependent, ß-funaltrexamine and nor-binaltorphimine-reversible manner. In vivo, P-317 (0.1 mg/kg, i.p. and 1 mg/kg, p.o.) inhibited GI transit, displayed a potent antinociceptive action in abdominal pain tests and did not influence the CNS. CONCLUSION: P-317 produced a potent analgesic and antidiarrhoeal action in the mouse GI tract after oral administration. Given lower expression of MOP and KOP mRNA in IBS-D patients, P-317 is a promising peptide-based drug candidate for IBS-D therapy.

Withaferin A inhibits experimental epithelial-mesenchymal transition in MCF-10A cells and suppresses vimentin protein level in vivo in breast tumors.

We have shown previously that withaferin A (WA), a bioactive component of the medicinal plant Withania somnifera, inhibits growth of cultured and xenografted human breast cancer cells and prevents breast cancer development and pulmonary metastasis incidence in a transgenic mouse model. The present study was undertaken to determine if the anticancer effect of WA involved inhibition of epithelial-mesenchymal transition (EMT). Experimental EMT induced by exposure of MCF-10A cells to tumor necrosis factor-α (TNF-α) and transforming growth factor-ß1 (TGF-ß) was partially reversed by treatment with WA but not by its structural analogs withanone or withanolide A. Combined TNF-α and TGF-ß treatments conferred partial protection against MCF-10A cell migration inhibition by WA. Inhibition of TNF-α and TGF-ß-induced MCF-10A cell migration by WA exposure was modestly attenuated by knockdown of E-cadherin protein. MCF-7 and MDA-MB-231 cells exposed to WA exhibited sustained (MCF-7) or transient (MDA-MB-231) induction of E-cadherin protein. On the other hand, the level of vimentin protein was increased markedly after 24 h treatment of MDA-MB-231 cells with WA. WA-induced apoptosis was not affected by vimentin protein knockdown in MDA-MB-231 cells. Protein level of vimentin was significantly lower in the MDA-MB-231 xenografts as well as in MMTV-neu tumors from WA-treated mice compared with controls. The major conclusions of the present study are that (a) WA treatment inhibits experimental EMT in MCF-10A cells, and (b) mammary cancer growth inhibition by WA administration is associated with suppression of vimentin protein expression in vivo.

Zinc and selenium levels in women with gestational diabetes mellitus at Medani Hospital, Sudan.

Gestational diabetes is a common medical disorder in pregnancy. There is a growing body of evidence of the association between zinc, selenium status and diabetes mellitus during pregnancy. A case-control study was conducted at Medani Hospital, Sudan, to compare zinc and selenium levels in pregnant women with gestational diabetes and normal pregnant women (controls). The two groups (31 in each arm) were well-matched in age, parity, gestational age, haemoglobin and body mass index. Zinc and selenium levels were measured using atomic absorption spectrophotometry. There were no significant differences in the median (interquartile) zinc (498.9 [395-703] vs 486.4 [404-667] µg/l, p = 0.905) and selenium (164.4 [61-415] vs 204 [68-541] µg/l, p = 0.838) values between the two groups. There were no significant correlations between zinc and selenium, or between these trace elements and body mass index, gestational age and blood glucose levels.

Anti-inflammatory and antinociceptive action of an orally available nociceptin receptor agonist SCH 221510 in a mouse model of inflammatory bowel diseases.

The nociceptin receptors (NOPs) are expressed in the gastrointestinal (GI) tract on muscle cell membranes and neurons, as well as the immune cells that infiltrate the mucosa. The involvement of NOPs in the pathophysiology of GI inflammation has been suggested, but due to the lack of selective NOP agonists, it never fully elucidated. Our aim was to characterize the anti-inflammatory and antinociceptive effect of the NOP agonist, SCH 221510 [3-endo-8-[bis(2-methylphenyl)methyl]-3-phenyl-8-azabicyclo [3.2.1]octan-3-ol], as a potential therapeutic strategy in the treatment of inflammatory bowel diseases (IBD). The anti-inflammatory action of SCH 221510 was determined after intraperitoneal, oral, and intracolonic administration of SCH 221510 (0.1-3.0 mg/kg once or twice daily) in mice treated with 2,4,6-trinitrobenzenesulfonic acid (TNBS). Antinociceptive action of SCH 221510 was evaluated in the mouse model of mustard oil (MO)-induced abdominal pain. Relative NOP mRNA expression was assessed in patients with IBD using real-time reverse transcriptase-polymerase chain reaction. We found that the expression of NOP mRNA was significantly decreased in patients with IBD. The administration (0.1 and 1.0 mg/kg i.p. twice daily and 3 mg/kg p.o. twice daily) of SCH 221510 attenuated TNBS colitis in mice. This effect was blocked by a selective NOP antagonist [J-113397 [(±)-1-[(3R*,4R*)-1-(cyclooctylmethyl)-3-(hydroxymethyl)-4-piperidinyl]-3-ethyl-1,3-dihydro-2H-benzimidazol-2-one]]. The intracolonic injections of SCH 221510 did not improve colitis in mice. The antinociceptive effect of SCH 221510 was observed after oral administration of SCH 221510 in MO-induced pain tests in mice with acute colitis. In conclusion, our results show a potent anti-inflammatory and antinociceptive effect upon selective activation of NOP receptors and suggest that the NOP agonist SCH 221510 is a promising drug candidate for future treatment of IBD.

Growth arrest by the antitumor steroidal lactone withaferin A in human breast cancer cells is associated with down-regulation and covalent binding at cysteine 303 of ß-tubulin.

Withaferin A (WA), a C5,C6-epoxy steroidal lactone derived from a medicinal plant (Withania somnifera), inhibits growth of human breast cancer cells in vitro and in vivo and prevents mammary cancer development in a transgenic mouse model. However, the mechanisms underlying the anticancer effect of WA are not fully understood. Herein, we report that tubulin is a novel target of WA-mediated growth arrest in human breast cancer cells. The G2 and mitotic arrest resulting from WA exposure in MCF-7, SUM159, and SK-BR-3 cells was associated with a marked decrease in protein levels of ß-tubulin. These effects were not observed with the naturally occurring C6,C7-epoxy analogs of WA (withanone and withanolide A). A non-tumorigenic normal mammary epithelial cell line (MCF-10A) was markedly more resistant to mitotic arrest by WA compared with breast cancer cells. Vehicle-treated control cells exhibited a normal bipolar spindle with chromosomes aligned along the metaphase plate. In contrast, WA treatment led to a severe disruption of normal spindle morphology. NMR analyses revealed that the A-ring enone in WA, but not in withanone or withanolide A, was highly reactive with cysteamine and rapidly succumbed to irreversible nucleophilic addition. Mass spectrometry demonstrated direct covalent binding of WA to Cys(303) of ß-tubulin in MCF-7 cells. Molecular docking indicated that the WA-binding pocket is located on the surface of ß-tubulin and characterized by a hydrophobic floor, a hydrophobic wall, and a charge-balanced hydrophilic entrance. These results provide novel insights into the mechanism of growth arrest by WA in breast cancer cells.

Withania somnifera root extract inhibits mammary cancer metastasis and epithelial to mesenchymal transition.

Though clinicians can predict which patients are at risk for developing metastases, traditional therapies often prove ineffective and metastatic disease is the primary cause of cancer patient death; therefore, there is a need to develop anti-metastatic therapies that can be administered over long durations to specifically inhibit the motility of cancer cells. Withaniasomnifera root extracts (WRE) have anti-proliferative activity and the active component, Withaferin A, inhibits the pro-metastatic protein, vimentin. Vimentin is an intermediate filament protein and is part of the epithelial to mesenchymal transition (EMT) program to promote metastasis. Here, we determined whether WRE standardized to Withaferin A (sWRE) possesses anti-metastatic activity and whether it inhibits cancer motility via inhibition of vimentin and the EMT program. Several formulations of sWRE were created to enrich for Withaferin A and a stock solution of sWRE in EtOH could recover over 90% of the Withaferin A found in the original extract powder. This sWRE formulation inhibited breast cancer cell motility and invasion at concentrations less than 1µM while having negligible cytotoxicity at this dose. sWRE treatment disrupted vimentin morphology in cell lines, confirming its vimentin inhibitory activity. To determine if sWRE inhibited EMT, TGF-ß was used to induce EMT in MCF10A human mammary epithelial cells. In this case, sWRE prevented EMT induction and inhibited 3-D spheroid invasion. These studies were taken into a human xenograft and mouse mammary carcinoma model. In both models, sWRE and Withaferin A showed dose-dependent inhibition of tumor growth and metastatic lung nodule formation with minimal systemic toxicity. Taken together, these data support the hypothesis that low concentrations of sWRE inhibit cancer metastasis potentially through EMT inhibition. Moreover, these doses of sWRE have nearly no toxicity in normal mouse organs, suggesting the potential for clinical use of orally administered WRE capsules.