Structure of phosphorylated-like RssB, the adaptor delivering σs to the ClpXP proteolytic machinery, reveals an interface switch for activation.

In enterobacteria such as Escherichia coli, the general stress response is mediated by σs, the stationary phase dissociable promoter specificity subunit of RNA polymerase. σs is degraded by ClpXP during active growth in a process dependent on the RssB adaptor, which is thought to be stimulated by the phosphorylation of a conserved aspartate in its N-terminal receiver domain. Here we present the crystal structure of full-length RssB bound to a beryllofluoride phosphomimic. Compared to the structure of RssB bound to the IraD anti-adaptor, our new RssB structure with bound beryllofluoride reveals conformational differences and coil-to-helix transitions in the C-terminal region of the RssB receiver domain and in the interdomain segmented helical linker. These are accompanied by masking of the α4-ß5-α5 (4-5-5) "signaling" face of the RssB receiver domain by its C-terminal domain. Critically, using hydrogen-deuterium exchange mass spectrometry, we identify σs-binding determinants on the 4-5-5 face, implying that this surface needs to be unmasked to effect an interdomain interface switch and enable full σs engagement and hand-off to ClpXP. In activated receiver domains, the 4-5-5 face is often the locus of intermolecular interactions, but its masking by intramolecular contacts upon phosphorylation is unusual, emphasizing that RssB is a response regulator that undergoes atypical regulation.

Early perturbation of Wnt signaling reveals patterning and invagination-evagination control points in molar tooth development.

Tooth formation requires complex signaling interactions both within the oral epithelium and between the epithelium and the underlying mesenchyme. Previous studies of the Wnt/ß-catenin pathway have shown that tooth formation is partly inhibited in loss-of-function mutants, and gain-of-function mutants have perturbed tooth morphology. However, the stage at which Wnt signaling is first important in tooth formation remains unclear. Here, using an Fgf8-promoter-driven, and therefore early, deletion of ß-catenin in mouse molar epithelium, we found that loss of Wnt/ß-catenin signaling completely deletes the molar tooth, demonstrating that this pathway is central to the earliest stages of tooth formation. Early expression of a dominant-active ß-catenin protein also perturbs tooth formation, producing a large domed evagination at early stages and supernumerary teeth later on. The early evaginations are associated with premature mesenchymal condensation marker, and are reduced by inhibition of condensation-associated collagen synthesis. We propose that invagination versus evagination morphogenesis is regulated by the relative timing of epithelial versus mesenchymal cell convergence regulated by canonical Wnt signaling. Together, these studies reveal new aspects of Wnt/ß-catenin signaling in tooth formation and in epithelial morphogenesis more broadly.

Telehepatology Satisfaction Is Associated with Ethnicity: The Real-World Experience of a Vulnerable Population with Fatty Liver Disease.

BACKGROUND: Since the coronavirus disease 2019 (COVID-19) pandemic began, telemedicine use has transformed healthcare delivery. Yet there is concern that telemedicine may widen care disparities for vulnerable populations, and patient experience data are limited. AIMS: We aimed to assess patient satisfaction with hepatology-related telemedicine (telehepatology) for delivery of fatty liver disease (FLD) care in a safety-net healthcare system. METHODS: Adult patients with FLD were surveyed regarding satisfaction with telehepatology. Clinical, demographic, resources, and social determinants of health (SDoH) data were collected to identify factors associated with satisfaction through multivariable modeling. RESULTS: From June 2020 to March 2022, 220 participants were enrolled: the median age was 52 years, 37% were men, and 68% were Hispanic. One hundred nineteen (54%) had prior telehepatology experience. Overall, satisfaction was high; 70% reported being somewhat or very satisfied. On univariate analysis, Hispanic ethnicity (versus non-Hispanic, OR 0.34, 95% CI 0.1-0.9, p = 0.03) and limited access to personal cellphone/internet (OR 0.16, 95% CI 0.04-0.6, p = 0.01) were associated with lower satisfaction. On multivariable logistic regression modeling adjusted for pandemic duration, age, sex, severity of liver disease, and coexisting liver disease, Hispanic ethnicity and lack of personal cellphone/internet remained independently associated with lower telehepatology satisfaction (OR 0.24, 95% CI 0.07-0.9, p = 0.03 and OR 0.2, 95% CI 0.04-0.9, p = 0.04, respectively). The association remained statistically significant after inclusion of various SDoH in the multivariable model. CONCLUSIONS: Satisfaction with telehepatology among FLD patients in a safety-net clinical setting was high overall. However, Hispanic ethnicity and lack of personal cellphone/internet were independently associated with lower telehepatology satisfaction. A better understanding of patients' experience with telehepatology is needed to identify reasons for dissatisfaction, and in-person visits should remain an option for patients to ensure equitable care.

Deciphering the mechanism and function of Hsp100 unfoldases from protein structure.

Hsp100 chaperones, also known as Clp proteins, constitute a family of ring-forming ATPases that differ in 3D structure and cellular function from other stress-inducible molecular chaperones. While the vast majority of ATP-dependent molecular chaperones promote the folding of either the nascent chain or a newly imported polypeptide to reach its native conformation, Hsp100 chaperones harness metabolic energy to perform the reverse and facilitate the unfolding of a misfolded polypeptide or protein aggregate. It is now known that inside cells and organelles, different Hsp100 members are involved in rescuing stress-damaged proteins from a previously aggregated state or in recycling polypeptides marked for degradation. Protein degradation is mediated by a barrel-shaped peptidase that physically associates with the Hsp100 hexamer to form a two-component system. Notable examples include the ClpA:ClpP (ClpAP) and ClpX:ClpP (ClpXP) proteases that resemble the ring-forming FtsH and Lon proteases, which unlike ClpAP and ClpXP, feature the ATP-binding and proteolytic domains in a single polypeptide chain. Recent advances in electron cryomicroscopy (cryoEM) together with single-molecule biophysical studies have now provided new mechanistic insight into the structure and function of this remarkable group of macromolecular machines.

Disentangling the impact of alcohol use and hepatitis C on insulin action in Latino individuals.

BACKGROUND: Alcohol, insulin resistance (IR), and hepatitis C (HCV) are all significant contributors to adverse outcomes of chronic liver disease. Latinos are disproportionately affected by these risk factors. We investigated the relationship between alcohol use and insulin action in a prospective cohort of Latino individuals with and without HCV. METHODS: One hundred fifty-three nondiabetic Latino individuals (60 HCV+, 93 HCV-) underwent clinical evaluation and metabolic testing; 56 had repeat testing over a median follow-up of 1.5 years. Peripheral IR and hepatic IR were measured via steady-state plasma glucose (SSPG) and endogenous glucose production during a two-step, 240-min insulin suppression test. Insulin secretion (IS) was measured using the graded glucose infusion test. Alcohol use was categorized as none, moderate (≤1 drink/day for women and ≤2 drinks/day for men), and heavy (>moderate). Multivariable models including HCV status assessed associations of alcohol use with baseline SSPG, hepatic IR and IS, and changes in these parameters over time. RESULTS: Overall, the median age was 44 years, 63.4% were male, 66.7% overweight/ obese, and 31.9% had heavy lifetime alcohol use while 60.4% had moderate lifetime alcohol use. SSPG and IS were similar by levels of alcohol use at baseline and alcohol use was not statistically significantly associated with change in these measures over time. However, lifetime daily heavy alcohol use (vs. not heavy, coef 2.4 µU-mg/kg-min-ml, p = 0.04) and HCV status (coef 4.4 µU-mg/kg-min-ml, p = 0.0003) were independently associated with higher baseline hepatic IR, and current heavy alcohol use was associated with greater change in hepatic IR in follow-up (coef 5.8 µU-mg/kg-min-ml, p = 0.03). CONCLUSIONS: In this cohort of Latino individuals, lifetime and current heavy alcohol use influenced hepatic IR and its change over time. Strategies to decrease rates of heavy alcohol use or increase abstinence along with lifestyle modification and anti-HCV therapy to reduce metabolic risk are critical to prevent adverse liver and metabolic outcomes in Latino individuals.

Evaluation of a pilot, community-led mental illness de-stigmatization theater intervention in rural uganda.

BACKGROUND: In rural areas of low- and middle- income countries, mental health care is often unavailable and inaccessible, and stigma is a major barrier to treatment. Destigmatization can increase treatment-seeking attitudes, community support, and acceptance of individuals suffering from mental illness. This study's primary objective was to evaluate the impact of a community-led, theater-based destigmatization campaign for mental illness conducted in the Busoga region of Eastern Uganda. METHODS: One hundred residents of the Busoga region were randomly selected via cluster sampling to complete a structured questionnaire assessing mental health stigma. Four focus groups were conducted for qualitative data on mental health stigma. Common misconceptions and specific points of stigma were identified from these responses, and local village health team personnel developed and performed a culturally-adapted theatrical performance addressing these points. Changes in perceptions of mental illness were measured among 57 attendees using two measures, the Broad Acceptance Scale (designed to reflect factors that contribute to structural stigma) and Personal Acceptance Scale (designed to reflect factors that contribute to interpersonal, or public stigma), before and after the performance. RESULTS: There was a significant increase in acceptance according to the Broad Acceptance Scale (p < .001) and Personal Acceptance Scale (p < .001). Qualitative responses from play attendees also indicated a decrease in stigma and an increased sense of the importance of seeking treatment for mentally ill patients. CONCLUSION: This study shows community-led, theater intervention may be an effective tool for the destigmatization of mental illness in rural areas of Uganda. Larger studies are needed to further test the efficacy of this approach and potential for longer-term scalabilityand sustainability.

Delivery of audiological diagnoses for infants: a linguistic analysis of clinical communication.

OBJECTIVE: To describe and analyse the linguistic structure of audiological diagnoses for infants, to determine ways to optimise the delivery of diagnostic information to parents during this typically emotive time. DESIGN: This study analysed the linguistic structure of audio-recorded infant diagnostic appointments. STUDY SAMPLE: Nine appointments conducted by four experienced paediatric audiologists were analysed. RESULTS: Diagnoses of normal hearing were delivered explicitly and in a straightforward manner. Positive aspects of this outcome were highlighted, and audiologists used the pronoun "we," conveying a feeling of teamwork. In contrast, when a hearing loss was diagnosed, the diagnosis included disfluencies and the use of hedging, although positive aspects were also emphasised. In these cases, audiologists used the pronoun "I," thereby taking ownership of the results. Differences in the topics raised by audiologists and parents highlighted a mis-match between the information provided and the information requested. Topics addressed by audiologists were primarily medical and procedural, whereas parents were concerned with causes, treatments and experiential information. CONCLUSIONS: The use of the above linguistic strategies may serve to minimise the significance and impact of the diagnosis. Whilst the data are unable to be generalised to other contexts, the study has generated in-depth and nuanced information about diagnosis delivery.

Total Neoadjuvant Therapy for Operable Pancreatic Cancer.

BACKGROUND: Overall survival (OS) for operable pancreatic cancer (PC) is optimized when 4-6 months of nonsurgical therapy is combined with pancreatectomy. Because surgery renders the delivery of postoperative therapy uncertain, total neoadjuvant therapy (TNT) is gaining popularity. METHODS: We performed a retrospective cohort study of patients with operable PC and compared TNT with shorter course neoadjuvant therapy (SNT). Primary outcomes of interest included completion of neoadjuvant therapy (NT) and resection of the primary tumor, receipt of 5 months of nonsurgical therapy, and median OS. RESULTS: We reviewed 541 consecutive patients from 2009 to 2019 including 226 (42%) with resectable PC and 315 (58%) with borderline resectable (BLR) PC. The median age was 66 years (IQR [59, 72]), and 260 (48%) patients were female. TNT was administered to 89 (16%) patients and SNT was administered to 452 (84%). Both groups were equally likely to complete intended NT and surgery (p = 0.90). Patients who received TNT and surgical resection were more likely to have a complete pathologic response (8% vs 4%, p < 0.01) and were more likely to receive at least 5 months of nonsurgical therapy (67% vs 45%, p < 0.01). The median OS was 26 months [IQR (15, 57)]; not reached among patients treated with TNT, and 25 months [IQR (15, 56)] among patients treated with SNT (p = 0.19). CONCLUSIONS: TNT ensures the delivery of intended systemic therapy prior to a complicated operation without decreasing the chance of successful surgery; a window of operability was not lost. Patients who can tolerate SNT will likely benefit from TNT.

Home Disposal Kits for Leftover Opioid Medications After Surgery: Do They Work?

BACKGROUND: Postoperative overprescribing is common, and many patients will have excess medications. An effective method to encourage disposal is lacking. We hypothesized that a convenient home disposal kit will result in more appropriate disposal of excess opioids. MATERIALS AND METHODS: We conducted a single-center prospective observational pilot study to evaluate the effectiveness of a postoperative opioid disposal kit. Patients in the intervention group received an opioid disposal kit and educational handout before discharge from the hospital. At the first follow-up visit, patients completed a survey in which they reported the remaining amount of pain medications from their original prescription and their plan for the excess medication. Patients were asked about risk factors for chronic opioid use. We used multivariable Poisson regression to identify independent factors associated with an increased likelihood of appropriate opioid disposal. RESULTS: The survey was offered to 904 patients with a response rate of 91.7%. After excluding those with missing data, 571 patients were included in the study. Overall, 83 (14.5%) patients never filled an opioid prescription, and 286 (60.0%) patients had tablets remaining at the time of the follow-up visit. Among those with tablets remaining, 52 received a home disposal kit, whereas 234 patients with tablets remaining did not. Patients who received the kit were more likely to dispose of opioid medications (54.9% versus 34.8%, relative risk = 1.8, 95% CI 1.3-2.5). No confounders were identified during multivariable analysis that increased a patient's likelihood of disposing excess medications. CONCLUSIONS: The provision of a convenient home disposal kit postoperatively increased patient-reported opioid disposal.

Unique sperm haplotypes are associated with phenotypically different sperm subpopulations in Astyanax fish.

BACKGROUND: The phenotypes of sperm are generally believed to be under the control of the diploid genotype of the male producing them rather than their own haploid genotypes, because developing spermatids share cytoplasm through intercellular bridges. This sharing is believed to homogenize their content of gene products. However, not all developing spermatids have identical gene products and estimates are that alleles at numerous gene loci are unequally expressed in sperm. This provides scope for the hypothesis that sperm phenotypes might be influenced by their unique haplotypes. Here we test a key prediction of this hypothesis. RESULTS: The haploid hypothesis predicts that phenotypically different sperm subpopulations should be genetically distinct. We tested this by genotyping different sperm subpopulations that were generated by exposing sperm to a chemical dye challenge (Hoechst 33342). Dye treatment caused the cells to swell and tend to clump together. The three subpopulations of sperm we distinguished in flow cytometry corresponded to single cells, and clumps of two or three. Cell clumping in the presence of the dye may reflect variation in cell adhesivity. We found that allelic contents differed among the three populations. Importantly, the subpopulations with clumped sperm cells were significantly enriched in allelic combinations that had previously been observed to have significantly lower transmission success. CONCLUSIONS: We show that at least one sperm phenotype is correlated with its haploid genotype. This supports a broader hypothesis that the haploid genotypes of sperm cells may influence their fitness, with potentially significant implications for the transmission of deleterious alleles or combinations of alleles to their offspring.

The role of sleep in adolescents' daily stress recovery: Negative affect spillover and positive affect bounce-back effects.

The present study examined the role of sleep in daily affective stress recovery processes in adolescents. Eighty-nine American adolescents recorded their emotions and stress through daily surveys and sleep with Fitbit devices for two weeks. Results show that objectively measured sleep (sleep onset latency and sleep debt) moderated negative affective responses to previous-day stress, such that stress-related negative affect spillover effects became more pronounced as amount of sleep decreased. Total sleep time and sleep debt moderated cross-day positive affect "bounce-back" effects. With more sleep, morning positive affect on days following high stress tended to bounce back to the levels that were common following low stress days. Conversely, if sleep was short following high stress days, positive affect remained low the next morning. No evidence for subjective sleep quality as a moderator of spillover/bounce-back effects was found. This research suggests that sleep quantity could relate to overnight affective stress recovery.

From snapshots to movies: Understanding early tooth development in four dimensions.

The developing tooth offers a model for the study of ectodermal appendage organogenesis. The signaling networks that regulate tooth development have been intensively investigated, but how cell biological responses to signaling pathways regulate dental morphogenesis remains an open question. The increasing use of ex vivo imaging techniques has enabled live tracking of cell behaviors over time in high resolution. While recent studies using these techniques have improved our understanding of tooth morphogenesis, important gaps remain that require additional investigation. In addition, some discrepancies have arisen between recent studies, and resolving these will advance our knowledge of tooth development. Developmental Dynamics 246:442-450, 2016. © 2017 Wiley Periodicals, Inc.

Statin Use and Risk of Cirrhosis and Related Complications in Patients With Chronic Liver Diseases: A Systematic Review and Meta-analysis.

BACKGROUND & AIMS: Statins have been variably shown to decrease risk and complications of chronic liver diseases (CLDs). We performed a systematic review and meta-analysis to evaluate the association between statins and risk of cirrhosis and related complications in patients with CLDs. METHODS: Through a systematic literature search up to March 2017, we identified 13 studies (3 randomized trials, 10 cohort studies) in adults with CLDs, reporting the association between statin use and risk of development of cirrhosis, decompensated cirrhosis, improvements in portal hypertension, or mortality. Pooled relative risk (RR) estimates with 95% confidence interval (CIs) were calculated using random effects model. Grading of Recommendations Assessment, Development and Evaluation criteria were used to assess quality of evidence. RESULTS: Among 121,058 patients with CLDs (84.5% with hepatitis C), 46% were exposed to statins. In patients with cirrhosis, statin use was associated with 46% lower risk of hepatic decompensation (4 studies; RR, 0.54; 95% CI, 0.46-0.62; I2 = 0%; moderate-quality evidence), and 46% lower mortality (5 studies; RR, 0.54; 95% CI, 0.47-0.61; I2 = 10%; moderate-quality evidence). In patients with CLD without cirrhosis, statin use was associated with a nonsignificant (58% lower) risk of development of cirrhosis or fibrosis progression (5 studies; RR, 0.42; 95% CI, 0.16-1.11; I2 = 99%; very-low-quality evidence). In 3 randomized controlled trials, statin use was associated with 27% lower risk of variceal bleeding or progression of portal hypertension (hazard ratio, 0.73; 95% CI, 0.59-0.91; I2 = 0%; moderate-quality evidence). CONCLUSIONS: Based on a systematic review and meta-analysis, statin use is probably associated with lower risk of hepatic decompensation and mortality, and might reduce portal hypertension, in patients with CLDs. Prospective observational studies and randomized controlled trials are needed to confirm this observation.

Greater intracortical inhibition associates with lower quadriceps voluntary activation in individuals with ACL reconstruction.

Decreased voluntary activation contributes to quadriceps weakness following anterior cruciate ligament reconstruction (ACLR). Alterations in neural excitability are likely responsible for reductions in quadriceps voluntary activation, and may be due to specific alterations in intracortical inhibition and facilitation. Therefore, we sought to determine if intracortical inhibition (SICI) and intracortical facilitation (ICF) associate with quadriceps voluntary activation in individuals with ACLR. Twenty-seven participants with a primary, unilateral ACLR were enrolled in this study. Bilateral central activation ratio (CAR) and paired-pulse transcranial magnetic stimulation were used to assess quadriceps voluntary activation, as well as SICI and ICF in the vastus medalis, respectively. Pearson Product Moment correlations were used to determine the association between CAR and (1) SICI, and (2) ICF in each limb. Lesser CAR associated with lesser SICI amplitude (r = 0.502, P = 0.008) in the ACLR limb. No associations in the contralateral limb were significant. Our results suggest greater intracortical inhibition associates with lesser voluntary activation in individuals with ACL. Implementing interventions that target intracortical inhibition may aid in restoring quadriceps voluntary activation following ACLR.

Mitotic stopwatch for the blast fungus Magnaporthe oryzae during invasion of rice cells.

To study nuclear dynamics of Magnaporthe oryzae, we developed a novel mitotic reporter strain with GFP-NLS (localized in nuclei during interphase but in the cytoplasm during mitosis) and H1-tdTomato (localized in nuclei throughout the cell cycle). Time-lapse confocal microscopy of the reporter strain during host cell invasion provided several new insights into nuclear division and migration in M. oryzae: (i) mitosis lasts about 5min; (ii) mitosis is semi-closed; (iii) septal pores are closed during mitosis; and (iv) a nucleus exhibits extreme constriction (approximately from 2µm to 0.5µm), elongation (over 5µm), and long migration (over 16µm). Our observations raise new questions about mechanisms controlling the mitotic dynamics, and the answers to these questions may result in new means to prevent fungal proliferation without negatively affecting the host cell cycle.

Utilizing Remote Real-Time Videoconferencing to Expand Access to Cancer Genetic Services in Community Practices: A Multicenter Feasibility Study.

BACKGROUND: Videoconferencing has been used to expand medical services to low-access populations and could increase access to genetic services at community sites where in-person visits with genetic providers are not available. OBJECTIVE: To evaluate the feasibility of, patient feedback of, and cognitive and affective responses to remote two-way videoconferencing (RVC) telegenetic services at multiple sociodemographically diverse community practices without access to genetic providers. METHODS: Patients at 3 community sites in 2 US states outside the host center completed RVC pretest (visit 1, V1) and post-test (visit 2, V2) genetic counseling for cancer susceptibility. Surveys evaluated patient experiences, knowledge, satisfaction with telegenetic and cancer genetics services, anxiety, depression, and cancer worry. RESULTS: A total of 82 out of 100 (82.0%) approached patients consented to RVC services. A total of 61 out of 82 patients (74%) completed pretest counseling and 41 out of 61 (67%) proceeded with testing and post-test counseling. A total of 4 out of 41 (10%) mutation carriers were identified: BRCA2, MSH2, and PMS2. Patients reported many advantages (eg, lower travel burden and convenience) and few disadvantages to RVC telegenetic services. Most patients reported feeling comfortable with the video camera--post-V1: 52/57 (91%); post-V2: 39/41 (95%)--and that their privacy was respected--post-V1: 56/57 (98%); post-V2: 40/41 (98%); however, some reported concerns that RVC might increase the risk of a confidentiality breach of their health information--post-V1: 14/57 (25%); post-V2: 12/41 (29%). While the majority of patients reported having no trouble seeing or hearing the genetic counselor--post-V1: 47/57 (82%); post-V2: 39/41 (95%)--51 out of 98 (52%) patients reported technical difficulties. Nonetheless, all patients reported being satisfied with genetic services. Compared to baseline, knowledge increased significantly after pretest counseling (+1.11 mean score, P=.005); satisfaction with telegenetic (+1.74 mean score, P=.02) and genetic services (+2.22 mean score, P=.001) increased after post-test counseling. General anxiety and depression decreased after pretest (-0.97 mean anxiety score, P=.003; -0.37 mean depression score, P=.046) and post-test counseling (-1.13 mean anxiety score, P=.003; -0.75 mean depression score, P=.01); state anxiety and cancer-specific worry did not significantly increase. CONCLUSIONS: Remote videoconferencing telegenetic services are feasible, identify genetic carriers in community practices, and are associated with high patient satisfaction and favorable cognitive and affective outcomes, suggesting an innovative delivery model for further study to improve access to genetic providers and services. Potential barriers to dissemination include technology costs, unclear billing and reimbursement, and state requirements for provider licensure.

Validating Constructs of the Depression, Anxiety, and Stress Scale-21 and Exploring Health Indicators to Predict the Psychological Outcomes of Students Enrolled in the Pacific Islands Cohort of College Students.

The Depression, Anxiety, and Stress Scale-21 (DASS-21) has been used in various countries to assess the mental states of individuals. The objectives of this study were to validate the DASS-21 for use in Guam, an island that endures a high burden of mental health challenges, such as suicide, and examine the predictive impact of selected health indicators on DASS-21 variables. Three years of data (2017-2019) were pooled from the Pacific Islands Cohort of College Students (PICCS) study conducted annually at the University of Guam. In total, 726 students were included in the secondary data analysis. MPlus statistical software was used to perform a confirmatory factor analysis (CFA) for the validation and structural equation modeling (SEM) for the predictive modeling. The results from the CFA suggested an acceptable model fit (RMSEA: 0.073, CFI: 0.901, TLI: 0.889, RMR: 0.044), while SEM suggested that sleep quality and physical activity were significant predictors of DASS-21 variables. Therefore, the DASS-21 is a valid instrument for measuring depression, anxiety, and stress among emerging adults in Guam.