Pannexin 3 deletion reduces fat accumulation and inflammation in a sex-specific manner.

BACKGROUND: Pannexin 3 (PANX3) is a channel-forming glycoprotein that enables nutrient-induced inflammation in vitro, and genetic linkage data suggest that it regulates body mass index. Here, we characterized inflammatory and metabolic parameters in global Panx3 knockout (KO) mice in the context of forced treadmill running (FEX) and high-fat diet (HFD). METHODS: C57BL/6N (WT) and KO mice were randomized to either a FEX running protocol or no running (SED) from 24 until 30 weeks of age. Body weight was measured biweekly, and body composition was measured at 24 and 30 weeks of age. Male WT and KO mice were fed a HFD from 12 to 28 weeks of age. Metabolic organs were analyzed for a panel of inflammatory markers and PANX3 expression. RESULTS: In females there were no significant differences in body composition between genotypes, which could be due to the lack of PANX3 expression in female white adipose tissue, while male KOs fed a chow diet had lower body weight and lower fat mass at 24 and 30 weeks of age, which was reduced to the same extent as 6 weeks of FEX in WT mice. In addition, male KO mice exhibited significantly lower expression of multiple pro-inflammatory genes in white adipose tissue compared to WT mice. While on a HFD body weight differences were insignificant, multiple inflammatory genes were significantly different in quadriceps muscle and white adipose tissue resulting in a more anti-inflammatory phenotype in KO mice compared to WT. The lower fat mass in male KO mice may be due to significantly fewer adipocytes in their subcutaneous fat compared to WT mice. Mechanistically, adipose stromal cells (ASCs) cultured from KO mice grow significantly slower than WT ASCs. CONCLUSION: PANX3 is expressed in male adult mouse adipose tissue and may regulate adipocyte numbers, influencing fat accumulation and inflammation.

Nucleotides released from palmitate-activated murine macrophages attract neutrophils.

Obesity and elevation of circulating free fatty acids are associated with an accumulation and proinflammatory polarization of macrophages within metabolically active tissues, such as adipose tissue, muscle, liver, and pancreas. Beyond macrophages, neutrophils also accumulate in adipose and muscle tissues during high-fat diets and contribute to a state of local inflammation and insulin resistance. However, the mechanisms by which neutrophils are recruited to these tissues are largely unknown. Here we used a cell culture system as proof of concept to show that, upon exposure to a saturated fatty acid, palmitate, macrophages release nucleotides that attract neutrophils. Moreover, we found that palmitate up-regulates pannexin-1 channels in macrophages that mediate the attraction of neutrophils, shown previously to allow transfer of nucleotides across membranes. These findings suggest that proinflammatory macrophages release nucleotides through pannexin-1, a process that may facilitate neutrophil recruitment into metabolic tissues during obesity.

Changes in hip flexor passive compliance do not account for improvement in vertical jump performance after hip flexor static stretching.

To date, there is limited research investigating stretching of antagonist muscles and its effects on agonist muscle function. The purpose of this research was to investigate the effects of pre-static stretching (pre-SS) of the hip flexor musculature on passive hip extension range of motion (ROM) and vertical jump height. Fifteen subjects reported to the laboratory on 4 separate days (D1, D2, D3, and D4). D1 was for familiarization, while on D2 to D4, subjects randomly completed 1 of 3 intervention conditions; no stretch (CON), hip flexor stretch (HFS), or hip extensor stretch (HES). Subject's pre- and post-intervention hip extension ROM were measured before performing 3 sets of pre- and post-maximal counter-movement vertical jumps. Vertical jump height was normalized to baseline for data analysis. A repeated-measures ANOVA with post hoc paired sample t-tests revealed a significant increase in vertical jump height in the HFS condition (1.74% ± 0.73; p ≤ 0.05) when compared with CON (-1.34% ± 0.96) or HES (-1.74% ± 0.65) conditions. There was also a significant increase in hip extension ROM after the HFS stretching protocol (6.5 ± 2.75%; p ≤ 0.05) when compared with the CON protocol (-1.73 ± 3.26); however, no significant difference when compared with the HES protocol (1.84 ± 2.79). A correlation analysis showed that the relative hip laxity of each subject had no effect on response to either condition nor did the magnitude of hip ROM change predict improvement in vertical jump. These results suggest that performing SS of the hip flexors may enhance vertical jump performance independent of changes in passive compliance of the hip flexor muscular tendon unit.

PANX3 Channels Regulate Architecture, Adhesion, Barrier Function, and Inflammation in the Skin.

The channel-forming glycoprotein PANX3 functions in cutaneous wound healing and keratinocyte differentiation, but its role in maintaining skin homeostasis through aging is not yet understood. We found that PANX3 is absent in newborn skin but becomes upregulated with age. We characterized the skin of global Panx3-knockout (KO) mice and found that KO dorsal skin showed sex differences at different ages but generally had reduced dermal and hypodermal areas compared with age-matched controls. Transcriptomic analysis of the KO epidermis revealed reduced E-cadherin stabilization and Wnt signaling compared with that of wild-type, consistent with the inability of primary KO keratinocytes to adhere in culture and diminished epidermal barrier function in KO mice. We also observed increased inflammatory signaling in the KO epidermis and a higher incidence of dermatitis in aged KO mice compared with that in wild-type controls. These findings suggest that during skin aging, PANX3 is critical in the maintenance of dorsal skin architecture, keratinocyte cell-cell and cell-matrix adhesion, and inflammatory skin responses.

Bone structure is largely unchanged in growing male CD-1 mice fed lower levels of vitamin D and calcium than in the AIN-93G diet.

BACKGROUND: Calcium (Ca) and vitamin D (vit D) in the AIN-93G diet may be higher than required for healthy bone development, and mask the potential benefit of a dietary intervention. OBJECTIVE: The objective was to determine if lower levels of Ca and vit D than is present in the AIN-93G diet supports bone development in growing male CD-1 mice. METHODS: Weanling male CD-1 mice were randomized to modified AIN-93G diets containing either 100 (Trial 1) or 400 (Trial 2) IU vit D/kg diet within one of two or three Ca levels (0.35, 0.30, or 0.25% Ca diet in Trial 1 or 0.35% or 0.25% in Trial 2) or the AIN-93G diet (1000 IU/kg vit D and 0.5% Ca) from weaning to 4 months of age (n = 13-15/group). At 2 and 4 months of age, BMD and structural properties of the tibia were analyzed in vivo. Structure of lumbar vertebra 4 (L4) and mandible, and femur strength were assessed ex vivo at age 4 months. RESULTS: There were no differences in tibia, L4, and mandible structure between the AIN-93G diet and the 0.35% Ca groups at either vit D level. A few structure outcomes were compromised with the 0.25 and/or 0.3% Ca diets but there were no differences in femur biomechanical strength compared to AIN-93G group in either Trial. CONCLUSION: At 400 or 100 IU vit D/kg diet, Ca can be lowered to 0.35% without detriment to BMD or bone structure while bone strength is not altered at lower Ca (0.25%) compared to CD-1 mice fed AIN-93G diet. Because of genetic variation in CD-1 mice among different breeding facilities, results in CD-1 mice from other facilities may differ from the present study.

Bone development in growing female mice fed calcium and vitamin D at lower levels than is present in the AIN-93G reference diet.

BACKGROUND: The AIN-93G reference (REF) diet is used to allow the comparison within and between studies of different research groups but its levels of vitamin D (vit D) and calcium (Ca) may be higher than required for healthy bone structure and bone mineral density (BMD). OBJECTIVE: To determine if lower dietary levels of Ca (3.5, 3 or 2.5 g Ca/kg diet) at 1 of 2 levels of vit D (100 or 400 IU/kg diet) supports similar development of bone structure and BMD compared to AIN-93G reference (REF) diet in female CD-1 mice at 2 and 4 months of age. METHODS: Within a trial, weanling female mice (n = 12-15/group) were randomized to 1 of 4 diets until necropsy at 4 months of age: Trial 1: 100 IU vit D/kg + 3.5, 3 or 2.5 g Ca/kg diet or 1000 IU vit D/kg + 5 g Ca/kg diet (REF); and Trial 2: 400 IU vit D/kg + 3.5, 3 or 2.5 g Ca/kg diet or 1000 IU vit D/kg + 5 g/kg diet (REF). At age 2 and 4 months, in vivo bone structure and BMD were assessed using micro-computed tomography (µCT) at the proximal and midpoint tibia. At age 4 months, lumbar vertebra 4 (L4) and mandible structure were analyzed ex vivo, femur strength at midpoint and neck was assessed and serum 25(OH)D3 and PTH were quantified. RESULTS: For Trial 1 (100 IU vit D/kg), there were no differences in tibia structure at age 2 and 4 months nor L4 or mandible structure or femur strength at the midpoint or neck at 4 months of age despite lower serum 25(OH)D3 among all groups compared to REF. For Trial 2 (400 IU vit D/kg), mice fed 2.5 g Ca/kg diet had lower (p < 0.05) Ct.Ar/Tt.Ar and Ct.Th at the tibia midpoint compared to REF. Furthermore, Ct.Th. was greater in REF and 3.5 g Ca/kg diet compared to 2.5 g Ca/kg diet at age 2 but not 4 months of age. At L4, BV/TV was lower (p < 0.05) in the 3 g Ca/kg diet group compared to REF at age 4 months. There were no differences among groups for serum 25(OH)D3 or femur strength at the midpoint or neck. Serum PTH was not elevated compared to REF in either Trial. CONCLUSION: Lowering both dietary vit D (100 IU/kg) and Ca (2.5 g/kg) in AIN-93G diet did not result in differences in bone development of female CD-1 mice at early adulthood. Translational relevance of bone studies conducted using the AIN-93G diet may be affected by its high vit D and Ca content.

Anti-Androgen Abiraterone Acetate Improves the Therapeutic Efficacy of Statins on Castration-Resistant Prostate Cancer Cells.

The treatment of castration-resistant (CR) prostate cancer (PCa) is limited. A sub-population of CR PCa tumors can synthesize androgens for intracrine androgen receptor (AR) activation, thus targeting androgen biosynthesis could be an effective therapeutic option for these patients. We determined that androgen biosynthesis inhibitors simvastatin, atorvastatin, and ketoconazole directly inhibit growth, migration, and colony formation of LNCaP C-81 cells, which exhibit de novo androgen biosynthesis, with simvastatin being the most effective. Importantly, in combination treatments, statins specifically enhanced growth suppression with added effects by anti-androgen abiraterone acetate on the CR PCa cells. Thus, statins can be used in conjunction with abiraterone acetate to enhance anti-androgen therapy for CR PCa.

Statin derivatives as therapeutic agents for castration-resistant prostate cancer.

Despite recent advances in modern medicine, castration-resistant prostate cancer remains an incurable disease. Subpopulations of prostate cancer cells develop castration-resistance by obtaining the complete steroidogenic ability to synthesize androgens from cholesterol. Statin derivatives, such as simvastatin, inhibit cholesterol biosynthesis and may reduce prostate cancer incidence as well as progression to advanced, metastatic phenotype. In this study, we demonstrate novel simvastatin-related molecules SVA, AM1, and AM2 suppress the tumorigenicity of prostate cancer cell lines including androgen receptor-positive LNCaP C-81 and VCaP as well as androgen receptor-negative PC-3 and DU145. This is achieved through inhibition of cell proliferation, colony formation, and migration as well as induction of S-phase cell-cycle arrest and apoptosis. While the compounds effectively block androgen receptor signaling, their mechanism of inhibition also includes suppression of the AKT pathway, in part, through disruption of the plasma membrane. SVA also possess an added effect on cell growth inhibition when combined with docetaxel. In summary, of the compounds studied, SVA is the most potent inhibitor of prostate cancer cell tumorigenicity, demonstrating its potential as a promising therapeutic agent for castration-resistant prostate cancer.

Reliability of goniometric and trigonometric techniques for measuring hip-extension range of motion using the modified Thomas test.

CONTEXT: Goniometric assessment of hip-extension range of motion is a standard practice in clinical rehabilitation settings. A weakness of goniometric measures is that small errors in landmarking may result in substantial measurement error. A less commonly used protocol for measuring hip range of motion involves applying trigonometric principles to the length and vertical displacement of the upper part of the lower extremity to determine hip angle; however, the reliability of this measure has never been assessed using the modified Thomas test. OBJECTIVE: To compare the intrarater and interrater reliability of goniometric (GON) and trigonometric (TRIG) techniques for assessing hip-extension range of motion during the modified Thomas test. DESIGN: Controlled laboratory study. SETTING: Institutional athletic therapy facility. PATIENTS OR OTHER PARTICIPANTS: A total of 22 individuals (12 men, 10 women; age range, 18-36 years) with no pathologic knee or back conditions. MAIN OUTCOME MEASURE(S): Hip-extension range of motion of each participant during a modified Thomas test was assessed by 2 examiners with both GON and TRIG techniques in a randomly selected order on 2 separate days. RESULTS: The intraclass correlation coefficient (ICC) revealed that the reliability of the GON technique was low for both the intrarater (ICC = 0.51, 0.54) and interrater (ICC = 0.30, 0.65) comparisons, but the reliability of the TRIG technique was high for both intrarater (ICC = 0.90, 0.95) and interrater (ICC = 0.91, 0.94) comparisons. Single-factorial repeated-measures analyses of variance revealed no mean differences in scoring within or between examiners for either measurement protocol, whereas a difference was observed when comparing the TRIG and GON tests due to the differences in procedures used to identify landmarks. CONCLUSIONS: Using the TRIG technique to measure hip-extension range of motion during the modified Thomas test results in superior intrarater and interrater reliability when compared with the GON technique.