Greater Protein Intake Emphasizing Lean Beef Does Not Affect Resistance Training-Induced Adaptations in Skeletal Muscle and Tendon of Older Women: A Randomized Controlled Feeding Trial.

BACKGROUND: Although experimental research supports that resistance training (RT), especially with greater dietary protein intake, improves muscle mass and strength in older adults, comparable research on tendons is needed. OBJECTIVES: We assessed the effects of a protein-rich diet emphasizing lean beef, compared with 2 control diets, on RT-induced changes in skeletal muscle and tendon size and strength in older women. METHODS: We randomly assigned women [age: 66 ± 1 y, body mass index (BMI): 28 ± 1] to groups that consumed 1) 0.8 g total protein/kg body weight/day from mixed food sources (normal protein control, n = 16); 2) 1.4 g/kg/d protein from mixed food sources (high protein control, n = 17); or 3) 1.4 g/kg/d protein emphasizing unprocessed lean beef (high protein experimental group, n = 16). Participants were provided with all foods and performed RT 3 times/wk, 70% of 1-repetition maximum for 12 wk. We measured quadriceps muscle volume via magnetic resonance imaging (MRI). We estimated patellar tendon biomechanical properties and cross-sectional area (CSA) using ultrasound and MRI. RESULTS: Dietary intake did not influence RT-induced increases in quadriceps strength (P < 0.0001) or muscle volume (P < 0.05). We noted a trend for an RT effect on mean tendon CSA (P = 0.07), with no differences among diets (P > 0.05). Proximal tendon CSA increased with RT (P < 0.05) with no difference between dietary groups (P > 0.05). Among all participants, midtendon CSA increased with RT (P ≤ 0.05). We found a decrease in distal CSA in the 0.8 g group (P < 0.05) but no change in the 1.4 g group (P > 0.05). Patellar tendon MRI signal or biomechanical properties were unchanged. CONCLUSIONS: Our findings indicated that greater daily protein intake, emphasizing beef, did not influence RT-induced changes in quadriceps muscle strength or muscle volume of older women. Although we noted trends in tendon CSA, we did not find a statistically significant impact of greater daily protein intake from beef on tendon outcomes. This trial was registered at clinicaltrials.gov as NCT04347447.

B2 cells contribute to hypertension and natural killer cell activation possibly via AT1-AA in response to placental ischemia.

Preeclampsia, new onset hypertension during pregnancy, is associated with activated T helper cells (Th) and B cells secreting agonistic autoantibodies against the angiotensin II type 1 receptor (AT1-AA). The reduced uterine perfusion pressure (RUPP) model of placental ischemia recapitulates these characteristics. We have shown that Th-B cell communication contributes to AT1-AA and symptoms of preeclampsia in the RUPP rat. B2 cells are classical B cells that communicate with Th cells and are then transformed into memory B cells. We hypothesize that B2 cells cause hypertension, natural killer (NK) cell activation, and complement activation during pregnancy through the production of AT1-AA. To test this hypothesis, total splenic B cells and B2 cells were isolated from normal pregnant (NP) or RUPP rats on gestational day (GD)19 and adoptively transferred into GD12 NP rats. A group of recipient rats was treated with a specific inhibitor peptide of AT1-AA. On GD19, mean arterial pressure was measured, tissues were collected, activated NK cells were measured by flow cytometry, and AT1-AA was measured by cardiomyocyte assay. NP recipients of RUPP B cells or RUPP B2 cells had increased mean arterial pressure, AT1-AA, and circulating activated NK cells compared with recipients of NP B cells. Hypertension in NP recipients of RUPP B cells or RUPP B2 was attenuated with AT1-AA blockade. This study demonstrates that B cells and B2 cells from RUPP rats cause hypertension and increased AT1-AA and NK cell activation in response to placental ischemia during pregnancy.NEW & NOTEWORTHY This study demonstrates that placental ischemia-stimulated B2 cells induce hypertension and circulating natural killer cell activation and angiotensin II type 1 receptor production in normal pregnant rats.

Impact of essential amino acid intake, resistance exercise, and aging on the concentration of Achilles peritendinous amino acids and procollagen Iα1 in humans.

Recent studies have shown that consuming amino acid-rich compounds improves tendon collagen content and biomechanical properties. Yet, it is unclear if the consumption of amino acids alters local (peritendinous) amino acid concentrations. If aging or exercise influence local amino acid concentrations in conjunction with an amino acid bolus is also not known. We conducted two studies. In Study 1, young women (n = 7, 25 ± 2 years) completed two identical resistance training sessions with either essential amino acid (EAA) or placebo consumption. In Study 2, an EAA bolus identical to Study 1 was given to younger (n = 7; 27 ± 1 year) and older adults (n = 6; 68 ± 2 years). Microdialysis was used to determine Achilles peritendinous amino acid and pro-collagen Iα1 (a marker of collagen synthesis) concentrations. In Study 1, amino acid consumption increased peritendinous concentrations of all EAA except histidine (p < 0.05). In Study 2, the peritendinous concentration of EAAs except for methionine, histidine, and lysine (p > 0.05) increased with time (p < 0.05). Further, the concentrations of most measured amino acids were greater in older adults (p < 0.05). Pro-collagen Iα1 concentration (p > 0.05) was unaffected by exercise, EAA, or aging (p > 0.05). Our findings demonstrate the following: (1) when not combined with exercise, an oral EAA bolus leads to only modest increases in Achilles peritendinous amino acid concentrations; (2) when combined with resistance exercise, EAA consumption resulted in greater peritendinous amino acid concentrations compared to no exercise; (3) the basal concentrations of most amino acids were greater in older adults, and (4) neither the EAA bolus nor exercise altered peritendinous pro-collagen concentrations.

Inhibition of the AT1R agonistic autoantibody in a rat model of preeclampsia improves fetal growth in late gestation.

Placenta ischemia, the initiating event in preeclampsia (PE), is associated with fetal growth restriction. Inhibition of the agonistic autoantibody against the angiotensin type 1 receptor AT1-AA, using an epitope-binding inhibitory peptide ('n7AAc') attenuates increased blood pressure at gestational day (G)19 in the clinically relevant reduced uterine perfusion pressure (RUPP) model of PE. Thus we tested the hypothesis that maternal administration of 'n7AAc' does not transfer to the fetus, improves uterine blood flow and fetal growth, and attenuates elevated placental expression of miRNAs implicated in PE and FGR. Sham or RUPP surgery was performed at G14 with vehicle or 'n7AAc' (144 µg/day) administered via an osmotic pump from G14 to G20. Maternal plasma levels of the peptide on G20 were 16.28 ± 4.4 nM, and fetal plasma levels were significantly lower at 1.15 ± 1.7 nM (P = 0.0007). The uterine artery resistance index was significantly elevated in RUPP (P < 0.0001) but was not increased in 'n7AAc'-RUPP or 'n7AAc'-Sham versus Sham. A significant reduction in fetal weight at G20 in RUPP (P = 0.003) was not observed in 'n7AAc'-RUPP. Yet, percent survival was reduced in RUPP (P = 0.0007) and 'n7AAc'-RUPP (P < 0.0002). Correlation analysis indicated the reduction in percent survival during gestation was specific to the RUPP (r = 0.5342, P = 0.043) and independent of 'n7AAc'. Placental miR-155 (P = 0.0091) and miR-181a (P = 0.0384) expression was upregulated in RUPP at G20 but was not elevated in 'n7AAc'-RUPP. Collectively, our results suggest that maternal administration of 'n7AAc' does not alter fetal growth in the RUPP implicating its potential as a therapeutic for the treatment of PE.NEW & NOTEWORTHY The seven amino acid inhibitory peptide to the AT1-AA ('n7AAc') has limited transfer to the fetus at gestational day 20, improves uterine blood flow and fetal growth in the reduced uterine perfusion pressure model of preeclampsia (PE), and does not impair fetal survival during gestation in sham-operated or placental ischemic rats. Collectively, these findings suggest that maternal administration of 'n7AAc' as an effective strategy for the treatment of PE is associated with improved outcomes in the fetus.

CD4+ T cells cause renal and placental mitochondrial oxidative stress as mechanisms of hypertension in response to placental ischemia.

The reduced uterine perfusion pressure (RUPP) rat model and normal pregnant (NP) rat recipients of RUPP CD4+ T cells recapitulate many characteristics of preeclampsia such as hypertension and oxidative stress. We have shown an important hypertensive role for natural killer (NK) cells to cause mitochondrial dysfunction in RUPP rats; however, the role for RUPP CD4+ T cells to stimulate NK cells is unknown. Therefore, we hypothesized that RUPP-induced CD4+ T cells activate NK cells to cause mitochondrial dysfunction/reactive oxygen species (ROS) as mechanisms of hypertension during pregnancy. We tested our hypothesis by adoptive transfer of RUPP CD4+ T cells into NP rats or by inhibiting the activation of RUPP CD4+ T cells with Orencia (abatacept) and examining hypertension, NK cells, and mitochondrial function. RUPP was performed on gestation day (GD) 14, and splenic CD4+ T cells were isolated on GD 19 and injected into NP rats on GD 13. In a separate group of rats, Orencia was infused and the RUPP procedure was performed. Mean arterial pressure and placental and renal mitochondrial ROS increased in RUPP (n = 7, P < 0.05) and NP + RUPP CD4+ T-cell recipients (n = 13, P < 0.05) compared with control NP (n = 7) and NP + NP CD4+ T-cell recipients (n = 5) but was reduced with Orencia (n = 13, P < 0.05). Placental and renal respiration was reduced in RUPP (n = 6, P < 0.05) and NP + RUPP CD4+ T-cell recipients (n = 6, state 3 P < 0.05) compared with NP (n = 5) and NP + NP CD4+ T-cell recipients (n = 5) but improved with Orencia (n = 9, n = 8 P < 0.05). These data indicate that CD4+ T cells, independent of NK cells, cause mitochondrial dysfunction/ROS contributing to hypertension in response to placental ischemia during pregnancy.

Open Reduction and Internal Fixation Versus Primary Arthrodesis for the Treatment of Acute Lisfranc Injuries: A Systematic Review and Meta-analysis.

This study aims to compare outcomes of open reduction and internal fixation (ORIF) and primary arthrodesis in management of Lisfranc injuries. In accordance with Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement standards, a systematic review was carried out. MEDLINE, EMBASE, CINAHL, and the Cochrane Central Register of Controlled Trials were searched to identify both randomised controlled trials (RCTs) and nonrandomised studies comparing the outcomes of ORIF and primary arthrodesis for Lisfranc injuries. Random- and fixed-effect statistical models were applied to calculate the pooled outcome data. Two RCTs and 3 observational studies were identified, compiling a total of 187 subjects with acute Lisfranc injuries and a mean follow-up duration of 62.3 months. Our results demonstrate that ORIF is associated with a significantly higher need for revision surgery (odds ratio [OR] 6.37, 95% confidence interval [CI] 2.68 to 15.11, p < .0001) and a significantly higher rate of persistent pain (OR 6.29, 95% CI 1.07 to 36.89, p = .04) compared with primary arthrodesis. However, we found no significant difference between the groups in terms of visual analogue scale pain score, American Orthopaedic Foot & Ankle Society functional score, or rates of infection. Separate analysis of RCTs showed that ORIF was associated with a more frequent need for revision surgery (OR 17.56, 95% CI 5.47 to 56.38, p < .00001), higher visual analogue scale pain score (mean difference 2.90, 95% CI 2.84 to 2.96, p < .00001), and lower American Orthopaedic Foot & Ankle Society score (mean difference -29.80, 95% CI -39.82 to -19.78, p < .00001). The results of the current study suggest that primary arthrodesis may be associated with better pain and functional outcomes and lower need for revision surgery compared with ORIF. The available evidence is limited and is not adequately robust to make explicit conclusions. The current literature requires high-quality and adequately powered RCTs.

Vitamin D supplementation reduces some AT1-AA-induced downstream targets implicated in preeclampsia including hypertension.

Autoantibodies to the ANG II type I receptor (AT1-AA) are associated with preeclampsia (PE). We found that vitamin D supplementation reduced AT1-AA and blood pressure (MAP) in the RUPP rat model of PE. However, it was undetermined whether the decrease in AT1-AA was the mechanism whereby vitamin D lowered MAP or if it were through factors downstream of AT1-AA. Uterine artery resistance index, placental ET-1, and soluble FMS-like tyrosine kinase-1 are increased with AT1-AA-induced hypertension and are considered markers of PE in pregnant women. Therefore, we hypothesized that vitamin D would reduce PE factors during AT1-AA-induced hypertension and could lower blood pressure in a model of hypertension during pregnancy without PE features. Either ANG II (50 ng·kg-1·day) or AT1-AA (1:40) was infused from gestational day (GD) 12-19. vitamin D2 (VD2, 270 IU/day) or vitamin D3 (VD3, 15 IU/day) was administered orally from GD14-GD18. MAP (mmHg) increased in AT1-AA (121 ± 4) and ANG II (113 ± 1)-infused pregnant rats compared with normal pregnant rats (NP) (101 ± 2) but was lower in AT1-AA+VD2 (105 ± 2), AT1-AA+VD3 (109 ± 2), ANG II+VD2 (104 ± 4), and ANG II+VD3 (104 ± 3). VD2 and/or VD3 improved PE features associated with AT1-AA during pregnancy, while ANG II did not induce such features, supporting the hypothesis that AT1-AA induces PE features during pregnancy, and these are improved with vitamin D. In this study, we demonstrate that vitamin D improved many factors associated with PE and reduced blood pressure in a hypertensive model without PE features, indicating that vitamin D could be beneficial for various hypertensive disorders of pregnancy.

Reduced uterine perfusion pressure T-helper 17 cells cause pathophysiology associated with preeclampsia during pregnancy.

Preeclampsia is associated with chronic inflammation and an imbalance among T-helper cell subtypes with an increase in T-helper 17 (TH17) cells. The objective of this study was to determine a role for TH17s, from the reduced uterine perfusion pressure (RUPP) rat model of preeclampsia, in the etiology of hypertension and chronic inflammation during pregnancy. CD4+/CD25- T cells were isolated from rat spleens, cultured in TH17 media, and were verified as TH17s via flow cytometry. On day 12 of gestation, 1×106 TH17 cells from RUPP rats were adoptively transferred into NP rats, carotid catheters were inserted on day 18, and on day 19, mean arterial pressure (MAP) was recorded, serum and plasma were collected, and oxidative stress and production of agonistic autoantibodies to the ANG II type I receptor (AT1-AA) were analyzed. MAP increased from 100.3 ± 1.7 mmHg in normal pregnant (NP; n = 17) to 124.8 ± 2.1 mmHg in RUPP (n = 22; P < 0.0001) and to 110.8 ± 2.8 mmHg in NP+RUPP TH17 (n = 11). Pup weights in NP+RUPP TH17s were decreased to 1.92 ± 0.09 g from 2.39 ± 0.14 in NP rats (P < 0.01). AT1-AA significantly increased from 0.1 ± 0.2 beats/min in NP to 15.6 ± 0.7 beats/min in NP+RUPP TH17s. IL-6 was 22.3 ± 5.7 pg/ml in NP and increased to 60.45 ± 13.8 pg/ml in RUPP (P < 0.05) and 75.9 ± 6.8 pg/ml in NP+RUPP TH17 rats (P < 0.01). Placental and renal oxidative stress were 238 ± 27.5 and 411 ± 129.9 relative light units·min-1·mg-1 in NP and 339 ± 104.6 and 833 ± 331.1 relative light units·min-1·mg-1 in NP+RUPP TH17, respectively. In conclusion, RUPP TH17 cells induced intrauterine growth restriction and increased blood pressure, AT1-AA, IL-6, and tissue oxidative stress when transferred to NP rats, indicating a role for autoimmune associated TH17 cells, to cause much of the pathophysiology associated with preeclampsia.

Transcriptomic response to heat stress among ecologically divergent populations of redband trout.

BACKGROUND: As ectothermic organisms have evolved to differing aquatic climates, the molecular basis of thermal adaptation is a key area of research. In this study, we tested for differential transcriptional response of ecologically divergent populations of redband trout (Oncorhynchus mykiss gairdneri) that have evolved in desert and montane climates. Each pure strain and their F1 cross were reared in a common garden environment and exposed over four weeks to diel water temperatures that were similar to those experienced in desert climates within the species' range. Gill tissues were collected from the three strains of fish (desert, montane, F1 crosses) at the peak of heat stress and tested for mRNA expression differences across the transcriptome with RNA-seq. RESULTS: Strong differences in transcriptomic response to heat stress were observed across strains confirming that fish from desert environments have evolved diverse mechanisms to cope with stressful environments. As expected, a large number of total transcripts (12,814) were differentially expressed in the study (FDR ≤ 0.05) with 2310 transcripts in common for all three strains, but the desert strain had a larger number of unique differentially expressed transcripts (2875) than the montane (1982) or the F1 (2355) strain. Strongly differentiated genes (>4 fold change and FDR ≤ 0.05) were particularly abundant in the desert strain (824 unique contigs) relative to the other two strains (montane = 58; F1 = 192). CONCLUSIONS: This study demonstrated patterns of acclimation (i.e., phenotypic plasticity) within strains and evolutionary adaptation among strains in numerous genes throughout the transcriptome. Key stress response genes such as molecular chaperones (i.e., heat shock proteins) had adaptive patterns of gene expression among strains, but also a much higher number of metabolic and cellular process genes were differentially expressed in the desert strain demonstrating these biological pathways are critical for thermal adaptation to warm aquatic climates. The results of this study further elucidate the molecular basis for thermal adaptation in aquatic ecosystems and extend the potential for identifying genes that may be critical for adaptation to changing climates.

Blockade of CD40 ligand for intercellular communication reduces hypertension, placental oxidative stress, and AT1-AA in response to adoptive transfer of CD4+ T lymphocytes from RUPP rats.

Preeclampsia (PE) is associated with altered immune activation during pregnancy. We have previously shown that adoptive transfer of CD4(+) T cells from the reduced uterine perfusion pressure (RUPP) rat model of PE increases blood pressure, oxidative stress (ROS), and inflammation in normal pregnant recipient rats. The objective of this study was to determine if blockade of communication via the CD40-CD40 ligand (CD40L) interaction between placental ischemia-induced CD4(+) T cells with endogenous normal pregnant (NP) cells would improve pathophysiology that was previously observed in NP recipient rats of RUPP CD4(+) T cells. Splenic CD4(+) T lymphocytes were magnetically separated, incubated with 2.5 µg/ml anti-CD40 ligand (αCD40L) overnight, and transferred into NP rats on day 12 of gestation (NP+RUPP CD4(+) T+anti-CD40L). On day 19 of gestation, blood pressure (MAP), blood, and tissues were collected. MAP was 99 ± 2 in NP (n = 13), 116 ± 4 in NP+RUPP CD4(+) T cells (n = 7; P < 0.01); MAP only increased to 104 ± 2 in NP+RUPP CD4(+) T cells+CD40L (n = 24) (P < 0.05 vs. NP+RUPP CD4(+) T cells). Mechanisms of hypertension in response to RUPP CD4(+) T cells include endothelin-1 (ET-1), ROS, and angiotensin II type I receptor (AT1-AA) were analyzed. Inhibition of CD40L binding reduced placental ET-1 to 2.3-fold above NP rats and normalized placental ROS from 318.6 ± 89 in NP+RUPP CD4(+) T cells (P < 0.05) to 118.7 ± 24 in NP+RUPP CD4(+) T+anti-CD40L (P < 0.05). AT1-AA was also normalized with inhibition of CD40L. These data suggest that placental ischemia-induced T-cell communication via the CD40L is one important mechanism leading to much of the pathophysiology of PE.

An increased population of regulatory T cells improves the pathophysiology of placental ischemia in a rat model of preeclampsia.

The reduced uterine perfusion pressure (RUPP) rat model of preeclampsia exhibits much of the pathology characterizing this disease, such as hypertension, inflammation, suppressed regulatory T cells (TRegs), reactive oxygen species (ROS), and autoantibodies to the ANG II type I receptor (AT1-AA) during pregnancy. The objective of this study was to determine whether supplementation of normal pregnant (NP) TRegs into RUPP rats would attenuate the pathophysiology associated with preeclampsia during pregnancy. CD4(+)/CD25(+) T cells were isolated from spleens of NP and RUPP rats, cultured, and injected into gestation day (GD) 12 normal pregnant rats that underwent the RUPP procedure on GD 14. On GD 1, mean arterial pressure (MAP) was recorded, and blood and tissues were collected for analysis. One-way ANOVA was used for statistical analysis. MAP increased from 99 ± 2 mmHg in NP (n = 12) to 127 ± 2 mmHg in RUPP (n = 21) but decreased to 118 ± 2 mmHg in RUPP+NP TRegs (n = 17). Circulating IL-6 and IL-10 were not significantly changed, while circulating TNF-α and IL-17 were significantly decreased after supplementation of TRegs. Placental and renal ROS were 339 ± 58.7 and 603 ± 88.1 RLU·min(-1)·mg(-1) in RUPP and significantly decreased to 178 ± 27.8 and 171 ± 55.6 RLU·min(-1)·mg(-1), respectively, in RUPP+NP TRegs; AT1-AA was 17.81 ± 1.1 beats per minute (bpm) in RUPP but was attenuated to 0.50 ± 0.3 bpm with NP TRegs. This study demonstrates that NP TRegs can significantly improve inflammatory mediators, such as IL-17, TNF-α, and AT1-AA, which have been shown to increase blood pressure during pregnancy.

The role of T cell stimulated agonistic autoantibodies to the angiotensin II type I receptor (AT1-AA) in mediating multiorgan dysfunction in IL-17 induced hypertension during pregnancy.

PROBLEM: Preeclampsia (PE), new-onset hypertension during pregnancy accompanied by organ dysfunction, is associated with chronic inflammation including elevated IL-17, CD4+ T cells, B cells and natural killer (NK) cells. IL-17 can serve as a signal for either the adaptive or innate immune activation. We have previously shown that IL-17 contributes to increased blood pressure in association with elevated TH17 cells, NK cells and B cells secreting angiotensin II type 1 receptor agonistic autoantibodies (AT1-AA) during pregnancy. Moreover, we have shown an important role for CD4+T cells and AT1-AA in multiorgan dysfunction as measured by mitochondrial oxidative stress (mt ROS). However, we do not know the role of adaptive immune cells such as T cells or B cells secreting AT1-AA in mediating the PE phenotype in response to elevated IL-17. METHOD OF STUDY: In order to answer this question, we infused IL-17 (150 pg/day i.p.) into either Sprague Dawley (SD) or athymic nude rats via mini-osmotic pump from gestational day (GD) 14-19 of pregnancy. On GD 19, blood pressure was determined and NK cells, mtROS and respiration and AT1-AA production from B cells were measured. RESULTS: Infusion of IL-17 increased blood pressure in the presence or absence of T cells. Mean arterial pressure (MAP) increased with IL-17 from 98 ± 2 mm Hg (n = 12) to 114 ± 2 (n = 12) in SD rats and from 99 ± 4 mm Hg (n = 7) versus 115 ± 2 mm Hg (n = 7) in athymic nude rats. Similar trends were seen in NK cells and placental mt ROS. Knowing that IL-17 stimulates AT1-AA in SD pregnant rats, we included a group of SD and athymic nude pregnant rats infused with IL-17 and the AT1-AA inhibitor peptide ('n7AAc'). The inhibitor attenuated blood pressure (104.9 ± 3.2, p = .0001) and normalized NK cells and mt function in SD pregnant rats. Importantly, the AT1-AA was not produced in pregnant nude IL-17 treated rats, nor did 'n7AAc' effect MAP, in nude athymic rats. CONCLUSION: These findings suggest two conclusions; one is that IL-17 causes hypertension and multiorgan dysfunction in the absence of T cells and AT1-AA, possibly through its activation of innate cells and secondly, in the presence of T cells, blockade of the AT1-AA attenuates the effect of IL-17. This study indicates the critical effects of elevated IL-17 during pregnancy and suggest treatment modalities to consider for PE women.

Development and validation of a GT-seq panel for genetic monitoring in a threatened species using minimally invasive sampling.

Minimally invasive samples are often the best option for collecting genetic material from species of conservation concern, but they perform poorly in many genomic sequencing methods due to their tendency to yield low DNA quality and quantity. Genotyping-in-thousands by sequencing (GT-seq) is a powerful amplicon sequencing method that can genotype large numbers of variable-quality samples at a standardized set of single nucleotide polymorphism (SNP) loci. Here, we develop, optimize, and validate a GT-seq panel for the federally threatened northern Idaho ground squirrel (Urocitellus brunneus) to provide a standardized approach for future genetic monitoring and assessment of recovery goals using minimally invasive samples. The optimized panel consists of 224 neutral and 81 putatively adaptive SNPs. DNA collected from buccal swabs from 2016 to 2020 had 73% genotyping success, while samples collected from hair from 2002 to 2006 had little to no DNA remaining and did not genotype successfully. We evaluated our GT-seq panel by measuring genotype discordance rates compared to RADseq and whole-genome sequencing. GT-seq and other sequencing methods had similar population diversity and F ST estimates, but GT-seq consistently called more heterozygotes than expected, resulting in negative F IS values at the population level. Genetic ancestry assignment was consistent when estimated with different sequencing methods and numbers of loci. Our GT-seq panel is an effective and efficient genotyping tool that will aid in the monitoring and recovery of this threatened species, and our results provide insights for applying GT-seq for minimally invasive DNA sampling techniques in other rare animals.

AT1-AA Is Produced in Offspring in Response to Placental Ischemia and Is Lowered by B-Cell Depletion Without Compromising Overall Offspring Health.

BACKGROUND: Preeclampsia, new-onset hypertension during pregnancy alongside other organ dysfunction, is the leading cause of mortality for the mother and low birth weight for the baby. Low birth weight contributes to high risk of cardiovascular disorders later in life. Women with preeclampsia have activated B cells producing agonistic autoantibodies to AT1-AA (angiotensin II type I receptor). We hypothesize that rituximab, a B cell-depleting chemotherapeutic, will deplete maternal B cells in reduced uterine perfusion pressure (RUPP) rats without worsening the effect of placental ischemia on pup growth and survival. METHODS AND RESULTS: To test this hypothesis, the RUPP procedure was performed, and rituximab was continuously infused via miniosmotic pump. Maternal blood and tissues were collected. A separate group of dams were allowed to deliver, pup weights were recorded, and at 4 months of age, tissues were collected from offspring. Immune cells were measured via flow cytometry, and AT1-AA was quantified using a contraction bioassay. Blood pressure increased in RUPP rats and was normalized with rituximab treatment. RUPP offspring also had increased circulating B cells, cytolytic natural killer cells, and increased circulating AT1-AA, which were normalized with maternal rituximab treatment. This is the first study to analyze the AT1-AA in RUPP offspring, which was normalized with rituximab. CONCLUSIONS: Our findings indicate that perinatal rituximab lowers maternal mean arterial pressure in RUPP rats and improves birth weight, circulating AT1-AA, and circulating natural killer cells, indicating that rituximab improves adverse fetal outcomes in response to placental ischemia.

Patellar tendon biomechanical and morphologic properties and their relationship to serum clinical variables in persons with prediabetes and type 2 diabetes.

Tendon biomechanical properties and fibril organization are altered in patients with diabetes compared to healthy individuals, yet few biomarkers have been associated with in vivo tendon properties. We investigated the relationships between in vivo imaging-based tendon properties, serum variables, and patient characteristics across healthy controls (n = 14, age: 45 ± 5 years, body mass index [BMI]: 24 ± 1, hemoglobin A1c [HbA1c]: 5.3 ± 0.1%), prediabetes (n = 14, age: 54 ± 5 years, BMI: 29 ± 2; HbA1c: 5.7 ± 0.1), and type 2 diabetes (n = 13, age: 55 ± 3 years, BMI: 33 ± 2, HbA1c: 6.7 ± 0.3). We used ultrasound speckle-tracking and measurements from magnetic resonance imaging (MRI) to estimate the patellar tendon in vivo tangent modulus. Analysis of plasma c-peptide, interleukin-1ß (IL-1ß), IL-6, IL-8, tumor necrosis factor-α (TNF-α), adiponectin, leptin, insulin-like growth factor 1 (IGF-1), and C-reactive protein (CRP) was completed. We built regression models incorporating statistically significant covariates and indicators for the clinically defined groups. We found that tendon cross-sectional area normalized to body weight (BWN CSA) and modulus were lower in patients with type 2 diabetes than in healthy controls (p < 0.05). Our regression analysis revealed that a model that included BMI, leptin, high-density lipoprotein (HDL), low-density lipoprotein (LDL), age, and group explained ~70% of the variability in BWN CSA (R2 = 0.70, p < 0.001). For modulus, including the main effects LDL, groups, HbA1c, age, BMI, cholesterol, IGF-1, c-peptide, leptin, and IL-6, accounted for ~54% of the variability in modulus (R2 = 0.54, p < 0.05). While BWN CSA and modulus were lower in those with diabetes, group was a poor predicter of tendon properties when considering the selected covariates. These data highlight the multifactorial nature of tendon changes with diabetes and suggest that blood variables could be reliable predictors of tendon properties.

Genome-wide association reveals genetic basis for the propensity to migrate in wild populations of rainbow and steelhead trout.

Little is known of the genetic basis of migration despite the ecological benefits migratory species provide to their communities and their rapid global decline due to anthropogenic disturbances in recent years. Using next-generation sequencing of restriction-site-associated DNA (RAD) tags, we genotyped thousands of single nucleotide polymorphisms (SNPs) in two wild populations of migratory steelhead and resident rainbow trout (Oncorhynchus mykiss) from the Pacific Northwest of the United States. One population maintains a connection to the sea, whereas the other population has been sequestered from its access to the ocean for more than 50 years by a hydropower dam. Here we performed a genome-wide association study to identify 504 RAD SNP markers from several genetic regions that were associated with the propensity to migrate both within and between the populations. Our results corroborate those in previous quantitative trait loci studies and provide evidence for additional loci associated with this complex migratory life history. Our results suggest a complex multi-genic basis with several loci of small effect distributed throughout the genome contributing to migration in this species. We also determined that despite being sequestered for decades, the landlocked population continues to harbour genetic variation associated with a migratory life history and ATPase activity. Furthermore, we demonstrate the utility of genotyping-by-sequencing and how RAD-tag SNP data can be readily compared between studies to investigate migration within this species.

Thermal adaptation and acclimation of ectotherms from differing aquatic climates.

To elucidate the mechanisms of thermal adaptation and acclimation in ectothermic aquatic organisms from differing climates, we used a common-garden experiment for thermal stress to investigate the heat shock response of redband trout (Oncorhynchus mykiss gairdneri) from desert and montane populations. Evidence for adaptation was observed as expression of heat shock genes in fish from the desert population was more similar to control (unstressed) fish and significantly different (P ≤ 0.05) from those from the montane population, while F1 crosses were intermediate. High induction of heat shock proteins (Hsps) in the montane strain appeared to improve short-term survival during first exposure to high water temperatures, but high physiological costs of Hsp production may have led to lower long-term survival. In contrast, the desert strain had significantly lower heat shock response than the montane fish and F1 crosses, suggesting that these desert fish have evolved alternative mechanisms to deal with thermal stress that provide better balance of physiological costs. Genomewide tests of greater than 10 000 SNPs found multiple SNPs that were significantly associated with survival under thermal stress, including Hsp47 which consistently appeared as a strong candidate gene for adaption to desert climates. Candidate SNPs identified in this study are prime targets to screen more broadly across this species' range to predict the potential for adaptation under scenarios of climate change. These results demonstrate that aquatic species can evolve adaptive responses to thermal stress and provide insight for understanding how climate change may impact ectotherms.

Population genomics of Pacific lamprey: adaptive variation in a highly dispersive species.

Unlike most anadromous fishes that have evolved strict homing behaviour, Pacific lamprey (Entosphenus tridentatus) seem to lack philopatry as evidenced by minimal population structure across the species range. Yet unexplained findings of within-region population genetic heterogeneity coupled with the morphological and behavioural diversity described for the species suggest that adaptive genetic variation underlying fitness traits may be responsible. We employed restriction site-associated DNA sequencing to genotype 4439 quality filtered single nucleotide polymorphism (SNP) loci for 518 individuals collected across a broad geographical area including British Columbia, Washington, Oregon and California. A subset of putatively neutral markers (N = 4068) identified a significant amount of variation among three broad populations: northern British Columbia, Columbia River/southern coast and 'dwarf' adults (F(CT) = 0.02, P ≪ 0.001). Additionally, 162 SNPs were identified as adaptive through outlier tests, and inclusion of these markers revealed a signal of adaptive variation related to geography and life history. The majority of the 162 adaptive SNPs were not independent and formed four groups of linked loci. Analyses with matsam software found that 42 of these outlier SNPs were significantly associated with geography, run timing and dwarf life history, and 27 of these 42 SNPs aligned with known genes or highly conserved genomic regions using the genome browser available for sea lamprey. This study provides both neutral and adaptive context for observed genetic divergence among collections and thus reconciles previous findings of population genetic heterogeneity within a species that displays extensive gene flow.

Sex Differences in Offspring of Preeclamptic Pregnancies.

A poor uterine environment causes changes in fetal development that affect the health of offspring long-term. Although there are multiple pathways that contribute to the development of cardiovascular and neurological disease, low birth weight or fetal growth restriction (FGR) predisposes offspring to these diseases. There is a link between fetal exposure to adverse influences and hypertension later in life. Many epidemiological studies support the link between fetal life and the risk of disease later in life. Experimental models have sought to provide mechanistic proof of this link while simultaneously investigating potential therapeutics or treatment pathways. Preeclampsia (PE), one of several hypertensive disorders in pregnancy, is a leading cause of morbidity and mortality for both the mother and fetus. Studies have shown that PE is a state of chronic inflammation and there is an imbalance between pro-inflammatory and regulatory immune cells and mediators. There is no cure for PE beyond the delivery of the fetal-placental unit, and many PE pregnancies result in FGR and preterm birth. Epidemiological data demonstrate that the sex of the offspring is correlated with the degree of cardiovascular disease that develops with the age of the offspring yet few studies examine the effect of sex on the development of neurological disorders. Even fewer studies examine the effects of therapeutics on offspring of different genders following a PE pregnancy. Moreover, there remain significant gaps in knowledge concerning the role the immune system plays in FGR offspring developing hypertension or neurovascular disorders later in life. Therefore, the purpose of this review is to highlight current research on sex differences in the developmental programming of hypertension and neurological disorders following a PE pregnancy.

The role of immune cells and mediators in preeclampsia.

Preeclampsia is a hypertensive disorder of major concern in pregnancy than can lead to intrauterine growth restriction, placental abruption and stillbirth. The pathophysiology of preeclampsia is multifactorial, including not only kidney dysfunction but also endothelial dysfunction, as the maternal endothelium becomes exposed to placental factors that are released into the circulation and increase systemic levels of vasoconstrictors, oxidative stress, anti-angiogenic factors and inflammatory mediators. Importantly, inflammation can lead to insufficient placental perfusion and low birthweight in offspring. Various innate and adaptive immune cells and mediators have been implicated in the development of preeclampsia, in which oxidative stress is associated with activation of the maternal inflammatory response. Immune cells such as regulatory T cells, macrophages, natural killer cells, and neutrophils are known to have major causative roles in the pathology of preeclampsia, but the contributions of additional immune cells such as B cells, inflammatory cytokines and anti-angiotensin II type 1 receptor autoantibodies are also now recognized. Immunological interventions, therefore, have therapeutic potential in this disease. Here, we provide an overview of the immune responses that are involved in the pathogenesis of preeclampsia, including the role of innate and adaptive immune cells and mediators.