Goats decrease hindlimb stiffness when walking over compliant surfaces.

Leg stiffness, commonly estimated as the 'compression' of a defined leg element in response to a load, has long been used to characterize terrestrial locomotion. This study investigated how goats adjust the stiffness of their hindlimbs to accommodate surfaces of different stiffness. Goats provide a compelling animal model for studying leg stiffness modulation, because they skillfully ambulate over a range of substrates that vary in compliance. To investigate the adjustments that goats make when walking over such substrates, ground reaction forces and three-dimensional trajectories of hindlimb markers were recorded as goats walked on rigid, rubber and foam surfaces. Net joint moments, power and work at the hip, knee, ankle and metatarsophalangeal joints were estimated throughout stance via inverse dynamics. Hindlimb stiffness was estimated from plots of total leg force versus total leg length, and individual joint stiffness was estimated from plots of joint moment versus joint angle. Our results support the hypothesis that goats modulate hindlimb stiffness in response to surface stiffness; specifically, hindlimb stiffness decreased on the more compliant surfaces (P<0.002). Estimates of joint stiffness identified hip and ankle muscles as the primary drivers of these adjustments. When humans run on compliant surfaces, they generally increase leg stiffness to preserve their center-of-mass mechanics. We did not estimate center-of-mass mechanics in this study; nevertheless, our estimates of hindlimb stiffness suggest that goats exhibit a different behavior. This study offers new insight into mechanisms that allow quadrupeds to modulate their gait mechanics when walking on surfaces of variable compliance.

The Assembled Genome of the Stroke-Prone Spontaneously Hypertensive Rat.

BACKGROUND: We report the creation and evaluation of a de novo assembly of the genome of the spontaneously hypertensive rat, the most widely used model of human cardiovascular disease. METHODS: The genome is assembled from long read sequencing (PacBio HiFi and continuous long read data [CLR]) and scaffolded with long-range structural information obtained from Bionano optical maps and proximity ligation sequencing proximity analysis of the genome. The genome assembly was polished with Illumina short reads. Completeness of the assembly was investigated using Benchmarking Universal Single Copy Orthologs analysis. The genome assembly was also evaluated with the rat reference gene set, using NCBI automated protocols. We also generated orthogonal single molecule transcript sequence reads (Iso-Seq) from 8 tissues and used them to validate the coding assembly, to annotate the assembly with RNA transcripts representing unique full length transcript isoforms for each gene and to determine whether divergences between RefSeq sequences and the assembly were attributable to assembly errors or polymorphisms. RESULTS: The assembly analysis indicates that this assembly is comparable in contiguity and completeness to the current rat reference assembly, while the use of HiFi sequencing yields an assembly that is more correct at the single base level. Synteny analysis was performed to uncover the extent of synteny and the presence and distribution of chromosomal rearrangements between the reference and this assembly. CONCLUSION: The resulting genome assembly is reference quality and captures significant structural variation.

Citrate concentrations in human seminal fluid and expressed prostatic fluid determined via 1H nuclear magnetic resonance spectroscopy outperform prostate specific antigen in prostate cancer detection.

PURPOSE: We compared the performance of citrate concentration measurements in unprocessed human semen and expressed prostatic secretions from controls and from patients with biopsy confirmed prostate cancer to that of prostate specific antigen testing with respect to specificity and sensitivity for prostate cancer detection. MATERIALS AND METHODS: Semen and expressed prostatic secretions were collected in biopsy proven, prostate cancer bearing and noncancer bearing cases. Citrate concentrations were determined by quantitative in vitro, high field, water suppressed proton nuclear magnetic resonance spectroscopy. Assessments of the diagnostic performance of citrate and prostate specific antigen results in our study populations were made by ROC curve analysis. RESULTS: Citrate was measured in samples from 61 participants, of whom 16 without and 21 with cancer donated semen, and 17 without and 7 with cancer donated expressed prostatic secretions. Mean citrate +/- SE compared to that in controls was 2.7-fold lower in patients with cancer samples in semen (132.2 +/- 30.1 vs 48.0 +/- 7.9 mM, p < 0.05) and expressed prostatic secretions (221.4 +/- 55.4 vs 81.5 +/- 36.0 mM, p < 0.05). ROC curve analysis showed that measurements of citrate in semen performed as well as measurements of citrate in expressed prostatic secretion for detecting prostate cancer (AUC 0.81, 95% CI 0.60 to 0.92 and AUC 0.73, 95% CI 0.38 to 0.90, respectively, p > 0.05). ROC curve analysis also showed that the measurement of citrate in either fluid outperformed prostate specific antigen measurement for detecting prostate cancer in these subjects (AUC 0.61, 95% CI 0.44 to 0.74). CONCLUSIONS: In vitro nuclear magnetic resonance spectroscopic measurement of the citrate concentration in semen or expressed prostatic secretions outperforms prostate specific antigen testing for detecting prostate cancer.

A decrease in 1H nuclear magnetic resonance spectroscopically determined citrate in human seminal fluid accompanies the development of prostate adenocarcinoma.

PURPOSE: Because human prostatic fluid contributes almost 50% of the volume of seminal plasma and this fluid contains unique prostatic metabolites such as citrate, which are markedly altered during tumorigenesis, we investigated high resolution H nuclear magnetic resonance (NMR) spectroscopy of unprocessed human seminal plasma as a rapid, noninvasive diagnostic tool for prostate adenocarcinoma. MATERIALS AND METHODS: Semen and prostatic massage samples from control and tumor bearing subjects were stored frozen at -20C and thawed prior to water suppressed NMR analysis. We found that freezing produced no significant alterations in the semen NMR spectra. Quantitative NMR spectroscopy was performed by first calibrating the water suppression data acquisition sequence with a series of standard samples containing known amounts of citrate within the physiological range. RESULTS: Well resolved citrate resonances from the seminal plasma of 3 control subjects with prostate specific antigen (PSA) less than 1 ng/ml were integrated to give concentrations of 97 to 178 mM. Semen from a 47-year-old man with benign prostatic hyperplasia and a PSA of 5.5 ng/ml contained 156 mM citrate. In contrast, seminal plasma from 2 patients with prostate cancer, including a 46-year-old man with Gleason grade 8 and PSA 45.2 ng/ml, and a 64-year-old man with grade 6 and PSA 13.0 ng/ml, revealed citrate NMR signals corresponding to a concentration of only 28 and 24 mM, respectively. Spectra from prostatic massage fluid from a normal 23-year-old volunteer showed a citrate of 483 mM, while massage fluid from a 56-year-old patient with Gleason grade 4 cancer showed a citrate of only 1.35 mM. CONCLUSIONS: To our knowledge this study is the first to use high resolution NMR of semen to diagnose prostate cancer. Given the known effects of adenocarcinoma on prostate metabolism, the study indicates that high resolution H NMR can be used to measure citrate in seminal fluid, potentially providing a new, rapid, noninvasive screening method.

Identification of residues at the alpha and epsilon subunit interfaces mediating species selectivity of Waglerin-1 for nicotinic acetylcholine receptors.

Waglerin-1 (Wtx-1) is a 22-amino acid peptide that is a competitive antagonist of the muscle nicotinic receptor (nAChR). We find that Wtx-1 binds 2100-fold more tightly to the alpha-epsilon than to the alpha-delta binding site interface of the mouse nAChR. Moreover, Wtx-1 binds 100-fold more tightly to the alpha-epsilon interface from mouse nAChR than that from rat or human sources. Site-directed mutagenesis of residues differing in the extracellular domains of rat and mouse epsilon subunits indicates that residues 59 and 115 mediate the species difference in Wtx-1 affinity. Mutation of residues 59 (Asp in mouse, Glu in rat epsilon) and 115 (Tyr in mouse, Ser in rat epsilon) converts Wtx-1 affinity for the alpha-epsilon interface of one species to that of the other species. Studies of different mutations at position 59 indicate both steric and electrostatic contributions to Wtx-1 affinity, whereas at position 115, both aromatic and polar groups contribute to affinity. The human nAChR also has lower affinity for Wtx-1 than mouse nAChR, but unlike rat nAChR, residues in both alpha and epsilon subunits mediate the affinity difference. In human nAChR, polar residues (Ser-187 and Thr-189) confer low affinity, whereas in mouse nAChR aromatic residues (Trp-187 and Phe-189) confer high affinity. The overall results show that non-conserved residues at the nAChR binding site, although not crucial for activation by ACh, govern the potency of neuromuscular toxins.