Long-duration leptin transgene expression in dorsal vagal complex does not alter bone parameters in female Sprague Dawley rats.

The hypothalamus and dorsal vagal complex (DVC) are both important for integration of signals that regulate energy balance. Increased leptin transgene expression in either the hypothalamus or DVC of female rats was shown to decrease white adipose tissue and circulating levels of leptin and adiponectin. However, in contrast to hypothalamus, leptin transgene expression in the DVC had no effect on food intake, circulating insulin, ghrelin and glucose, nor on thermogenic energy expenditure. These findings imply different roles for hypothalamus and DVC in leptin signaling. Leptin signaling is required for normal bone accrual and turnover. Leptin transgene expression in the hypothalamus normalized the skeletal phenotype of leptin-deficient ob/ob mice but had no long-duration (≥10 weeks) effects on the skeleton of leptin-replete rats. The goal of this investigation was to determine the long-duration effects of leptin transgene expression in the DVC on the skeleton of leptin-replete rats. To accomplish this goal, we analyzed bone from three-month-old female rats that were microinjected with recombinant adeno-associated virus encoding either rat leptin (rAAV-Leptin, n = 6) or green fluorescent protein (rAAV-GFP, control, n = 5) gene. Representative bones from the appendicular (femur) and axial (3rd lumbar vertebra) skeleton were evaluated following 10 weeks of treatment. Selectively increasing leptin transgene expression in the DVC had no effect on femur cortical or cancellous bone microarchitecture. Additionally, increasing leptin transgene expression had no effect on vertebral osteoblast-lined or osteoclast-lined bone perimeter or marrow adiposity. Taken together, the findings suggest that activation of leptin receptors in the DVC has minimal specific effects on the skeleton of leptin-replete female rats.

Six months of voluntary alcohol consumption in male cynomolgus macaques reduces intracortical bone porosity without altering mineralization or mechanical properties.

Chronic heavy alcohol consumption is a risk factor for low trauma bone fracture. Using a non-human primate model of voluntary alcohol consumption, we investigated the effects of 6 months of ethanol intake on cortical bone in cynomolgus macaques (Macaca fascicularis). Young adult (6.4 ± 0.1 years old, mean ± SE) male cynomolgus macaques (n = 17) were subjected to a 4-month graded ethanol induction period, followed by voluntary self-administration of water or ethanol (4 % w/v) for 22 h/d, 7 d/wk. for 6 months. Control animals (n = 6) consumed an isocaloric maltose-dextrin solution. Tibial response was evaluated using densitometry, microcomputed tomography, histomorphometry, biomechanical testing, and Raman spectroscopy. Global bone response was evaluated using biochemical markers of bone turnover. Monkeys in the ethanol group consumed an average of 2.3 ± 0.2 g/kg/d ethanol resulting in a blood ethanol concentration of 90 ± 12 mg/dl in longitudinal samples taken 7 h after the daily session began. Ethanol consumption had no effect on tibia length, mass, density, mechanical properties, or mineralization (p > 0.642). However, compared to controls, ethanol intake resulted in a dose-dependent reduction in intracortical bone porosity (Spearman rank correlation = -0.770; p < 0.0001) and compared to baseline, a strong tendency (p = 0.058) for lower plasma CTX, a biochemical marker of global bone resorption. These findings are important because suppressed cortical bone remodeling can result in a decrease in bone quality. In conclusion, intracortical bone porosity was reduced to subnormal values 6 months following initiation of voluntary ethanol consumption but other measures of tibia architecture, mineralization, or mechanics were not altered.

Bone Marrow Adipose Tissue Is Not Required for Reconstitution of the Immune System Following Irradiation in Male Mice.

Bone marrow adipose tissue (BMAT) is hypothesized to serve as an expandable/contractible fat depot which functions, in part, to minimize energy requirements for sustaining optimal hematopoiesis. We investigated whether BMAT is required for immune reconstitution following injury. Male wild type (WBB6F1, WT) and BMAT-deficient WBB6F1/J-KitW/KitW-v/J (KitW/W-v) mice were lethally irradiated. Irradiation was followed by adoptive transfer of 1000 purified WT hematopoietic stem cells (HSCs). The extent of immune reconstitution in blood, bone marrow, and lymph nodes in the irradiated mice was determined using HSCs from green fluorescent protein (GFP)-expressing mice. We also evaluated skeletal response to treatment. Detection of GFP-positive B and T cells in peripheral blood at 4 and 9 weeks following adoptive transfer and in bone marrow and lymph nodes following necropsy revealed excellent immune reconstitution in both WT and BMAT-deficient mice. Adipocytes were numerous in the distal femur of WT mice but absent or rare in KitW/W-v mice. Bone parameters, including length, mass, density, bone volume, microarchitecture, and turnover balance, exhibited few differences between WT and BMAT-deficient mice. The minimal differences suggest that BMAT is not required for reconstitution of the immune system following lethal radiation and is not a major contributor to the skeletal phenotypes of kit signaling-deficient mice.

Higher weight in partially leptin-resistant db/+ mice is associated with positive effects on bone.

Absence of leptin confers metabolic dysfunction resulting in morbid obesity. Bone growth and maturation are also impaired. Partial leptin resistance is more common than leptin deficiency and, when induced by feeding mice a high fat diet, often has a negative effect on bone. Here, we used a genetic model to investigate the skeletal effects of partial and total leptin resistance in mice. This was accomplished by comparing the skeletal phenotypes of 17-week-old female C57Bl6/J wild-type (WT) mice, partial leptin receptor-deficient (db/+) mice and leptin receptor-deficient (db/db) mice (n = 7-8/group), all fed a standard diet. Compared to WT mice, db/db mice were dramatically heavier and hyperleptinemic. These mice were also hypogonadal, hyperglycemic, osteopenic and had lower serum levels of bone turnover markers, osteocalcin and C-terminal telopeptide of type I collagen (CTX). Compared to WT mice, db/+ mice were 14% heavier, had 149% more abdominal white adipose tissue, and were mildly hyperglycemic. db/+ mice did not differ from WT mice in uterine weight or serum levels of markers of bone turnover, although there was a trend for lower osteocalcin. At the bone microarchitectural level, db/+ mice differed from WT mice in having more massive femurs and a trend (P = 0.072) for larger vertebrae. These findings suggest that db/+ mice fed a normal mouse diet compensate for partial leptin resistance by increasing white adipose tissue mass which results in higher leptin levels. Our findings suggest that db/+ mice are a useful diet-independent model for studying the effects of partial leptin resistance on the skeleton.

Risk Factors for Salmonella Detection in Commercial Layer Flocks in Spain.

Trends in Salmonella human infections are assumed to be related to the distribution of the pathogen in the animal reservoir/food products, and cases in humans are most often linked to poultry and poultry products (eggs, meat). Therefore, ongoing Salmonella national control programs (NCPs) in European Union Member States have the objective of monitoring and reducing its prevalence in commercial poultry flocks. Results from NCPs have shown certain factors (housing systems, season of sampling and if sampling is conducted by food business operators (FBOps) or competent authorities (CAs), among others) can influence detection rates, but associations are often not consistent. Here, we analyzed data from the Spanish NCP on 7216 laying hen flocks subjected to 36,193 sampling events over a six-year period to characterize its performance and identify variables influencing detection rates. Overall, 1205 sampling events were positive for Salmonella spp. (any serovar) and 132 for S. Enteritidis-S. Typhimurium/monophasic. Bayesian multivariable models adjusting for multiple covariates concluded that sampling events later in the year, in caged flocks with older animals and conducted by CAs had increased odds of positivity for Salmonella spp., revealing aspects linked with a differential estimation of Salmonella levels in laying hen flocks.

Diet composition influences the effect of high fat diets on bone in growing male mice.

The effect of diet-induced obesity on bone in rodents is variable, with bone mass increases, decreases, and no impact reported. The goal of this study was to evaluate whether the composition of obesogenic diet may influence bone independent of its effect on body weight. As proof-of-principle, we used a mouse model to compare the skeletal effects of a commonly used high fat 'Western' diet and a modified high fat diet. The modified high fat diet included ground English walnut and was isocaloric for macronutrients, but differed in fatty acid composition and contained nutrients (e.g. polyphenols) not present in the standard 'Western' diet. Eight-week-old mice were randomized into 1 of 3 dietary treatments (n = 8/group): (1) low fat control diet (LF; 10 % kcal fat); (2) high fat 'Western' diet (HF; 46 % kcal fat as soybean oil and lard); or (3) modified high fat diet supplemented with ground walnuts (HF + walnut; 46 % kcal fat as soybean oil, lard, and walnut) and maintained on their respective diets for 9 weeks. Bone response in femur was then evaluated using dual energy x-ray absorptiometry, microcomputed tomography, and histomorphometry. Consumption of both obesogenic diets resulted in increased weight gain but differed in impact on bone and bone marrow adiposity in distal femur metaphysis. Mice consuming the high fat 'Western' diet exhibited a tendency for lower cancellous bone volume fraction and connectivity density, and had lower osteoblast-lined bone perimeter (an index of bone formation) and higher bone marrow adiposity than low fat controls. Mice fed the modified high fat diet did not differ from mice fed control (low fat) diet in cancellous bone microarchitecture, or osteoblast-lined bone perimeter, and exhibited lower bone marrow adiposity compared to mice fed the 'Western' diet. This proof-of-principal study demonstrates that two obesogenic diets, similar in macronutrient distribution and induction of weight gain, can have different effects on cancellous bone in distal femur metaphysis. Because the composition of the diets used to induce obesity in rodents does not recapitulate a common human diet, our finding challenges the translatability of rodent studies evaluating the impact of diet-induced obesity on bone.

Narrative minireview of the spatial epidemiology of substance use disorder in the United States: Who is at risk and where?

Drug overdose is the leading cause of death by injury in the United States. The incidence of substance use disorder (SUD) in the United States has increased steadily over the past two decades, becoming a major public health problem for the country. The drivers of the SUD epidemic in the United States have changed over time, characterized by an initial heroin outbreak between 1970 and 1999, followed by a painkiller outbreak, and finally by an ongoing synthetic opioid outbreak. The nature and sources of these abused substances reveal striking differences in the socioeconomic and behavioral factors that shape the drug epidemic. Moreover, the geospatial distribution of the SUD epidemic is not homogeneous. The United States has specific locations where vulnerable communities at high risk of SUD are concentrated, reaffirming the multifactorial socioeconomic nature of this epidemic. A better understanding of the SUD epidemic under a spatial epidemiology framework is necessary to determine the factors that have shaped its spread and how these patterns can be used to predict new outbreaks and create effective mitigation policies. This narrative minireview summarizes the current records of the spatial distribution of the SUD epidemic in the United States across different periods, revealing some spatiotemporal patterns that have preceded the occurrence of outbreaks. By analyzing the epidemic of SUD-related deaths, we also describe the epidemic behavior in areas with high incidence of cases. Finally, we describe public health interventions that can be effective for demographic groups, and we discuss future challenges in the study and control of the SUD epidemic in the country.

Small changes in thermoregulation influence cancellous bone turnover balance in distal femur metaphysis in growing female mice.

Mice are typically housed at temperatures well below their thermoneutral zone. When individually housed at room temperature (~22 °C) mice experience cold stress which results in cancellous bone loss and has the potential to alter the skeletal response to treatment. It is not clear if there is a threshold temperature for cold stress-induced bone loss. It is also not clear if alternative strategies for attenuating cold stress, such as group housing, influence bone accrual and turnover. This study aimed to determine how small differences in temperature (4 °C) or heat loss (individual versus group housing with nestlets) influence bone in growing female C57BL/6 J mice. Five-week-old mice were randomized by weight to 1 of 4 treatment groups (N = 10/group): 1) baseline, 2) single housed at 22 °C, 3) single housed at 26 °C, or 4) group housed (n = 5/cage) with nestlets at 22 °C. Mice in the baseline group were sacrificed 1 week later, at 6 weeks of age. The other 3 groups of mice were maintained at their respective temperatures and housing conditions for 13 weeks until 18 weeks of age. Compared to baseline, mice single housed at room temperature had increased body weight and femur size, but dramatically decreased cancellous bone volume fraction in distal femur metaphysis. The cancellous bone loss was attenuated but not prevented in mice individually housed at 26 °C or group housed at 22 °C. In conclusion, by impacting thermogenesis or heat loss, modest differences in housing conditions could influence experimental results.

Cold stress during room temperature housing alters skeletal response to simulated microgravity (hindlimb unloading) in growing female C57BL6 mice.

Laboratory mice are typically housed at temperatures below the thermoneutral zone for the species, resulting in cold stress and premature cancellous bone loss. Furthermore, mice are more dependent upon non-shivering thermogenesis to maintain body temperature during spaceflight, suggesting that microgravity-induced bone loss may be due, in part, to altered thermogenesis. Consequently, we assessed whether housing mice at room temperature modifies the skeletal response to simulated microgravity. This possibility was tested using the hindlimb unloading (HLU) model to mechanically unload femora. Humeri were also assessed as they remain weight bearing during HLU. Six-week-old female C57BL6 (B6) mice were housed at room temperature (22 °C) or near thermoneutral (32 °C) and HLU for 2 weeks. Compared to baseline, HLU resulted in cortical bone loss in femur, but the magnitude of reduction was greater in mice housed at 22 °C. Cancellous osteopenia in distal femur (metaphysis and epiphysis) was noted in HLU mice housed at both temperatures. However, bone loss occurred at 22 °C, whereas the bone deficit at 32 °C was due to failure to accrue bone. HLU resulted in cortical and cancellous bone deficits (compared to baseline) in humeri of mice housed at 22 °C. In contrast, fewer osteopenic changes were detected in mice housed at 32 °C. These findings support the hypothesis that environmental temperature alters the skeletal response to HLU in growing female mice in a bone compartment-specific manner. Taken together, species differences in thermoregulation should be taken into consideration when interpreting the skeletal response to simulated microgravity.

Sanitary sewage overflows, boil water advisories, and emergency room and urgent care visits for gastrointestinal illness: a case-crossover study in South Carolina, USA, 2013-2017.

BACKGROUND: Sanitary sewage overflows (SSOs) release raw sewage, which may contaminate the drinking water supply. Boil water advisories (BWAs) are issued during low or negative pressure events, alerting customers to potential contamination in the drinking water distribution system. OBJECTIVE: We evaluated the associations between SSOs and BWAs and diagnoses of gastrointestinal (GI) illness in Columbia, South Carolina, and neighboring communities, 2013-2017. METHODS: A symmetric bi-directional case-crossover study design was used to assess the role of SSOs and BWAs on Emergency Room and Urgent Care visits with a primary diagnosis of GI illness. Cases were considered exposed if an SSO or BWA occurred 0-4 days, 5-9 days, or 10-14 days prior to the diagnosis, within the same residential zip code. Effect modification was explored via stratification on participant-level factors (e.g., sex, race, age) and season (January-March versus April-December). RESULTS: There were 830 SSOs, 423 BWAs, and 25,969 cases of GI illness. Highest numbers of SSOs, BWAs and GI cases were observed in a zip code where >80% of residents identified as Black or African-American. SSOs were associated with a 13% increase in the odds of a diagnosis for GI illness during the 0-4 day hazard period, compared to control periods (Odds Ratio: 1.13, 95% Confidence Interval: 1.09, 1.18), while no associations were observed during the other hazard periods. BWAs were not associated with increased or decreased odds of GI illness during all three hazard periods. However, in stratified analyses BWAs issued between January-March were associated with higher odds of GI illness, compared to advisories issued between April-December, in all three hazard periods. SIGNIFICANCE: SSOs (all months) and BWAs (January-March) were associated with increased odds of a diagnosis of GI illness. Future research should examine sewage contamination of the drinking water distribution system, and mechanisms of sewage intrusion from SSOs. IMPACT: Sewage contains pathogens, which cause gastrointestinal (GI) illness. In Columbia, South Carolina, USA, between 2013-2017, there were 830 sanitary sewage overflows (SSOs). There were also 423 boil water advisories, which were issued during negative pressure events. Using case-crossover design, SSOs (all months) and boil water advisories (January-March) were associated with increased odds of Emergency Room and Urgent Care diagnoses of GI illness, potentially due to contamination of the drinking water distribution system. Lastly, we identified a community where >80% of residents identified as Black or African-American, which experienced a disproportionate burden of sewage exposure, compared to the rest of Columbia.

Severe Zinc Deficiency Impairs Accrual of Bone in Rapidly Growing Rats That Is Partially Corrected Following Short-term Zinc Repletion.

Zinc (Zn) deficiency impairs bone growth. However, the precise skeletal effects of varying levels of Zn deficiency and response to subsequent Zn repletion on the growing skeleton are incompletely understood. To address this gap in knowledge, we investigated the effects of dietary Zn ((severe deficiency (< 0.5 mg Zn/kg diet) and short-term Zn repletion (30 mg/kg diet), marginal deficiency (6 mg Zn/kg diet)) on bone mass, density, and cortical and cancellous bone microarchitecture in growing male Sprague Dawley rats. Marginal Zn intake for 42 days had no effect on bone mass or cortical and cancellous bone microarchitecture. Twenty-one days of severe Zn deficiency lowered serum osteocalcin and C terminal telopeptide of type I collagen (CTX-1), decreased tibial bone mineral content and density, and lowered cross-sectional volume, cortical volume, and cortical thickness in tibial diaphysis as compared to both Zn-adequate (30 mg/kg diet) and pair-fed controls. Severe Zn deficiency similarly lowered cancellous bone volume in proximal tibial metaphysis. Zn repletion (10 days) accelerated weight gain, indicative of catch-up growth, normalized CTX-1 and osteocalcin, but did not normalize bone mass (unadjusted and adjusted for body weight) or cortical and cancellous bone microarchitecture. In summary, severe but not marginal Zn deficiency in rapidly growing rats impaired acquisition of cortical and cancellous bone, resulting in abnormalities in bone microarchitecture. Zn repletion accelerated weight gain compared to Zn-adequate controls but absence of a compensatory increase in serum osteocalcin or bone mass suggests Zn repletion may be insufficient to fully counteract the detrimental effects of prior Zn deficiency on skeletal growth.

Ethanol alters the relationship between IGF-1 and bone turnover in male macaques.

Insulin-like growth factor 1 (IGF-1) influences bone turnover. Transient decreases in IGF-I levels and/or bioavailability may contribute to the detrimental effects of alcohol on bone. The goals of this non-human primate study were to i) evaluate the 20-h response of bone turnover markers to ethanol consumption and ii) assess how ethanol consumption influences the relationship between IGF-1 and these markers. Osteocalcin (bone formation), carboxyterminal cross-linking telopeptide of type 1 collagen (CTX, bone resorption), IGF-1, and IGF binding protein 1 (IGFBP-1) were measured in plasma from male rhesus macaques (N = 10, 8.4 ± 0.3 years) obtained at 12:00, 16:00, and 06:00 h during two phases: pre-ethanol (alcohol-naïve) and ethanol access. During the ethanol access phase, monkeys consumed 1.5 g/kg/day ethanol (4% w/v) beginning at 10:00 h. Osteocalcin and CTX were lower, and the ratio of osteocalcin to CTX was higher at each time point during ethanol access compared to the pre-ethanol phase. Pre-ethanol marker levels did not vary across time points, but markers varied during ethanol access. IGF-1 levels, but not IGFBP-1 levels, varied during the pre-ethanol phase. In contrast, IGF-1 levels were stable during ethanol access but IGFBP-1 levels varied. There were positive relationships between IGF-1 and turnover markers during the pre-ethanol phase, but not during ethanol access. In conclusion, chronic ethanol consumption reduces levels of bone turnover markers and blocks the normal positive relationship between IGF-1 and turnover markers and alters the normal relationship between IGF-1 and IGFBP-1. These findings support the hypothesis that chronic alcohol consumption leads to growth hormone/IGF-1 resistance.

Leptin and environmental temperature as determinants of bone marrow adiposity in female mice.

Bone marrow adipose tissue (BMAT) levels are higher in distal femur metaphysis of female mice housed at thermoneutral (32°C) than in mice housed at 22°C, as are abdominal white adipose tissue (WAT) mass, and serum leptin levels. We performed two experiments to explore the role of increased leptin in temperature-enhanced accrual of BMAT. First, we supplemented 6-week-old female C57BL/6J (B6) mice with leptin for 2 weeks at 10 µg/d using a subcutaneously implanted osmotic pump. Controls consisted of ad libitum (ad lib) fed mice and mice pair fed to match food intake of leptin-supplemented mice. The mice were maintained at 32°C for the duration of treatment. At necropsy, serum leptin in leptin-supplemented mice did not differ from ad lib mice, suggesting suppression of endogenous leptin production. In support, Ucp1 expression in BAT, percent body fat, and abdominal WAT mass were lower in leptin-supplemented mice. Leptin-supplemented mice also had lower BMAT and higher bone formation in distal femur metaphysis compared to the ad lib group, changes not replicated by pair-feeding. In the second experiment, BMAT response was evaluated in 6-week-old female B6 wild type (WT), leptin-deficient ob/ob and leptin-treated (0.3 µg/d) ob/ob mice housed at 32°C for the 2-week duration of the treatment. Compared to mice sacrificed at baseline (22°C), BMAT increased in ob/ob mice as well as WT mice, indicating a leptin independent response to increased temperature. However, infusion of ob/ob mice with leptin, at a dose rate having negligible effects on either energy metabolism or serum leptin levels, attenuated the increase in BMAT. In summary, increased housing temperature and increased leptin have independent but opposing effects on BMAT in mice.

Isoliquiritigenin Decreases Bone Resorption and Osteoclast Differentiation.

SCOPE: A dose-ranging study is performed using young estrogen-depleted rats to determine whether dietary isoliquiritigenin (ILQ) alters bone metabolism and if the effects are associated with estrogen receptor signaling. METHODS AND RESULTS: Six-week-old rats (ovariectomized at 4 weeks of age) are fed diets containing 0, 100, 250, or 750 ppm ILQ (n = 5/treatment) for 7 days. Gene expression in femur and uterus, blood markers of bone turnover, body composition, and uterine weight and epithelial cell height are determined. Because ILQ lowers bone resorption, the effect of ILQ on in vitro differentiation of osteoclasts from bone marrow of mice is assessed. Treatment resulted in a dose-dependent increases in serum ILQ but no changes in serum osteocalcin, a marker of global bone formation. Contrastingly, ILQ administration results in reduced serum CTX-1, a marker of global bone resorption, and reduces tartrate resistant acid phosphatase expression in osteoclast culture. ILQ treatment and endogenous estrogen production had limited overlap on gene expression in femur and uterus. However, uterine epithelial cell hyperplasia is observed in two of five animals treated with 750 ppm. CONCLUSIONS: In conclusion, dietary ILQ reduces bone resorption in vivo and osteoclast differentiation in vitro, by mechanisms likely differing from actions of ovarian hormones.

Between-subject and within-subject variability in measures of biochemical markers of bone turnover in cynomolgus and rhesus macaques.

Development of optimal bone mass during early adulthood is determined by the balance between bone formation and resorption. The utility of minimally invasive biomarkers for monitoring bone turnover balance in maturing non-human primates has received limited attention. This study evaluated the biological variation of osteocalcin (a marker of bone formation), carboxyterminal cross-linking telopeptide of type 1 collagen (CTX, a marker of bone resorption), and the ratio of osteocalcin to CTX (reflecting bone turnover balance), in 136 rhesus and cynomolgus macaques aged 3.8-11.6 years. In a subsample of the animals (n = 28), blood samples were collected at monthly intervals over 4 months. Between-subject analysis revealed that there were no sex or species differences for CTX. Osteocalcin and the ratio of osteocalcin to CTX were higher in males than in females, and in rhesus macaques than in cynomolgus macaques. There were no changes in osteocalcin, CTX, or the ratio of osteocalcin to CTX across 4 months for any of the groups. In contrast, there was considerable within-subject variation in osteocalcin and CTX concentrations. However, differences in values exhibited no discernible pattern, suggesting that within-subject variation can be reduced by averaging repeat measurements. In summary, the data provide reference values for male and female rhesus and cynomolgus macaques and support the utility of osteocalcin and CTX as biomarkers to monitor bone turnover at the population level.

Rapamycin impairs bone accrual in young adult mice independent of Nrf2.

Advanced age is the strongest risk factor for osteoporosis. The immunomodulator drug rapamycin extends lifespan in numerous experimental model organisms and is being investigated as a potential therapeutic to slow human aging, but little is known about the effects of rapamycin on bone. We evaluated the impact of rapamycin treatment on bone mass, architecture, and indices of bone turnover in healthy adult (16-20 weeks old at treatment initiation) female wild-type (ICR) and Nrf2-/- mice, a mouse model of oxidative damage and aging-related disease vulnerability. Rapamycin (4 mg/kg bodyweight) was administered by intraperitoneal injection every other day for 12 weeks. Mice treated with rapamycin exhibited lower femur bone mineral content, bone mineral density, and bone volume compared to vehicle-treated mice. In midshaft femur diaphysis (cortical bone), rapamycin-treated mice had lower cortical volume and thickness, and in the distal femur metaphysis (cancellous bone), rapamycin-treated mice had higher trabecular spacing and lower connectivity density. Mice treated with rapamycin exhibited lower bone volume, bone volume fraction, and trabecular thickness in the 5th lumbar vertebra. Rapamycin-treated mice had lower levels of bone formation in the distal femur metaphysis compared to vehicle-treated mice which occurred co-incidentally with increased serum CTX-1, a marker of global bone resorption. Rapamycin had no impact on tibia inflammatory cytokine gene expression, and we found no independent effects of Nrf2 knockout on bone, nor did we find any interactions between genotype and treatment. These data show that rapamycin may have a negative impact on the skeleton of adult mice that should not be overlooked in the clinical context of its usage as a therapy to retard aging and reduce the incidence of age-related pathologies.

Caloric Restriction and Hypothalamic Leptin Gene Therapy Have Differential Effects on Energy Partitioning in Adult Female Rats.

Dieting is a common but often ineffective long-term strategy for preventing weight gain. Similar to humans, adult rats exhibit progressive weight gain. The adipokine leptin regulates appetite and energy expenditure but hyperleptinemia is associated with leptin resistance. Here, we compared the effects of increasing leptin levels in the hypothalamus using gene therapy with conventional caloric restriction on weight gain, food consumption, serum leptin and adiponectin levels, white adipose tissue, marrow adipose tissue, and bone in nine-month-old female Sprague-Dawley rats. Rats (n = 16) were implanted with a cannula in the 3rd ventricle of the hypothalamus and injected with a recombinant adeno-associated virus, encoding the rat gene for leptin (rAAV-Lep), and maintained on standard rat chow for 18 weeks. A second group (n = 15) was calorically-restricted to match the weight of the rAAV-Lep group. Both approaches prevented weight gain, and no differences in bone were detected. However, calorically-restricted rats consumed 15% less food and had lower brown adipose tissue Ucp-1 mRNA expression than rAAV-Lep rats. Additionally, calorically-restricted rats had higher abdominal white adipose tissue mass, higher serum leptin and adiponectin levels, and higher marrow adiposity. Caloric restriction and hypothalamic leptin gene therapy, while equally effective in preventing weight gain, differ in their effects on energy intake, energy expenditure, adipokine levels, and body composition.

"Know your epidemic, know your response": Epidemiological assessment of the substance use disorder crisis in the United States.

The United States (U.S.) is currently experiencing a substance use disorders (SUD) crisis with an unprecedented magnitude. The objective of this study was to recognize and characterize the most vulnerable populations at high risk of SUD mortality in the U.S., and to identify the locations where these vulnerable population are located. We obtained the most recent available mortality data for the U.S. population aged 15-84 (2005-2017) from the Centers for Diseases and Prevention (CDC). Our analysis focused on the unintentional substance poisoning to estimate SUD mortality. We computed health-related comorbidities and socioeconomic association with the SUD distribution. We identified the most affected populations and conducted a geographical clustering analysis to identify places with increased concentration of SUD related deaths. From 2005-2017, 463,717 SUD-related deaths occurred in the United States. White population was identified with the highest SUD death proportions. However, there was a surge of the SUD epidemic in the Black male population, with a sharp increase in the SUD-related death rate since 2014. We also found that an additional average day of mental distress might increase the relative risk of SUD-related mortality by 39%. The geographical distribution of the epidemic showed clustering in the West and Mid-west regions of the U.S. In conclusion, we found that the SUD epidemic in the U.S. is characterized by the emergence of several micro-epidemics of different intensities across demographic groups and locations within the country. The comprehensive description of the epidemic presented in this study could assist in the design and implementation of targeted policy interventions for addiction mitigation campaigns.

Identification of Vulnerable Populations and Areas at Higher Risk of COVID-19-Related Mortality during the Early Stage of the Epidemic in the United States.

We characterized vulnerable populations located in areas at higher risk of COVID-19-related mortality and low critical healthcare capacity during the early stage of the epidemic in the United States. We analyze data obtained from a Johns Hopkins University COVID-19 database to assess the county-level spatial variation of COVID-19-related mortality risk during the early stage of the epidemic in relation to health determinants and health infrastructure. Overall, we identified highly populated and polluted areas, regional air hub areas, race minorities (non-white population), and Hispanic or Latino population with an increased risk of COVID-19-related death during the first phase of the epidemic. The 10 highest COVID-19 mortality risk areas in highly populated counties had on average a lower proportion of white population (48.0%) and higher proportions of black population (18.7%) and other races (33.3%) compared to the national averages of 83.0%, 9.1%, and 7.9%, respectively. The Hispanic and Latino population proportion was higher in these 10 counties (29.3%, compared to the national average of 9.3%). Counties with major air hubs had a 31% increase in mortality risk compared to counties with no airport connectivity. Sixty-eight percent of the counties with high COVID-19-related mortality risk also had lower critical care capacity than the national average. The disparity in health and environmental risk factors might have exacerbated the COVID-19-related mortality risk in vulnerable groups during the early stage of the epidemic.

Dynamics of the COVID-19 epidemic in urban and rural areas in the United States.

PURPOSE: There is a growing concern about the COVID-19 epidemic intensifying in rural areas in the United States (U.S.). In this study, we described the dynamics of COVID-19 cases and deaths in rural and urban counties in the U.S. METHODS: Using data from April 1 to November 12, 2020, from Johns Hopkins University, we estimated COVID-19 incidence and mortality rates and conducted comparisons between urban and rural areas in three time periods at the national level, and in states with higher and lower COVID-19 incidence rates. RESULTS: Results at the national level showed greater COVID-19 incidence rates in urban compared to rural counties in the Northeast and Mid-Atlantic regions of the U.S. at the beginning of the epidemic. However, the intensity of the epidemic has shifted to a rapid surge in rural areas. In particular, high incidence states located in the Mid-west of the country had more than 3,400 COVID-19 cases per 100,000 people compared to 1,284 cases per 100,000 people in urban counties nationwide during the third period (August 30 to November 12). CONCLUSIONS: Overall, the current epicenter of the epidemic is located in states with higher infection rates and mortality in rural areas. Infection prevention and control efforts including healthcare capacity should be scaled up in these vulnerable rural areas.