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The human pharmacokinetics, metabolism, and excretion of [ 14 C]-ganaxolone (GNX) were characterized in healthy male subjects ( n = 8) following a single 300-mg (150 μ Ci) oral dose. GNX exhibited a short half-life of 4 hours in plasma, whereas total radioactivity had a half-life of 413 hours indicating extensive metabolism to long-lived metabolites. Identification of the major GNX circulating metabolites required extensive isolation and purification for liquid chromatography-tandem mass spectrometry analysis, together with in vitro studies, NMR spectroscopy, and synthetic chemistry support. This revealed that the major routes of GNX metabolism involved hydroxylation at the 16 α -hydroxy position, stereoselective reduction of the 20-ketone to afford the corresponding 20 α -hydroxysterol, and sulfation of the 3 α -hydroxy group. This latter reaction yielded an unstable tertiary sulfate, which eliminated the elements of H 2 SO 4 to introduce a double bond in the A ring. A combination of these pathways, together with oxidation of the 3 β -methyl substituent to a carboxylic acid and sulfation at the 20 α position, led to the major circulating metabolites in plasma, termed M2 and M17. These studies, which led to the complete or partial identification of no less than 59 metabolites of GNX, demonstrated the high complexity of the metabolic fate of this drug in humans and demonstrated that the major circulating products in plasma can result from multiple sequential processes that may not be easily replicated in animals or with animal or human in vitro systems. SIGNIFICANCE STATEMENT: Studies on the metabolism of [ 14 C]-ganaxolone in humans revealed a complex array of products that circulated in plasma, the two major components of which were formed via an unexpected multi-step pathway. Complete structural characterization of these (disproportionate) human metabolites required extensive in vitro studies, along with contemporary mass spectrometry, NMR spectroscopy, and synthetic chemistry efforts, which served to underscore the limitations of traditional animal studies in predicting major circulating metabolites in man., (Copyright © 2023 by The American Society for Pharmacology and Experimental Therapeutics.)

Insulin-like growth factor binding protein 4 (IGFBP4) is involved in adipogenesis, and IGFBP4 null mice have decreased body fat through decreased PPAR-γ expression. In the current study, we assessed whether variation in the IGFBP4 coding region influences body mass index (BMI) in American Indians who are disproportionately affected by obesity. Whole exome sequence data from a population-based sample of 6779 American Indians with longitudinal measures of BMI were used to identify variation in IGFBP4 that associated with BMI. A novel variant that predicts a p.Ser76Thr in IGFBP4 (Thr-allele frequency = 0.02) was identified which associated with the maximum BMI measured during adulthood (BMI 39.8 kg/m 2 for Thr-allele homozygotes combined with heterozygotes vs. 36.2 kg/m 2 for Ser-allele homozygotes, β = 6.7% per Thr-allele, p = 8.0 × 10 -5 , adjusted for age, sex, birth-year and the first five genetic principal components) and the maximum age- and sex-adjusted BMI z-score measured during childhood/adolescence (z-score 0.70 SD for Thr-allele heterozygotes vs. 0.32 SD for Ser-allele homozygotes, β = 0.37 SD per Thr-allele, p = 8.8 × 10 -6 ). In vitro functional studies showed that IGFBP4 with the Thr-allele (BMI-increasing) had a 55% decrease (p = 0.0007) in FOXO-induced transcriptional activity, reflecting increased activation of the PI3K/AKT pathway mediated through increased IGF signaling. Over-expression and knock-down of IGFBP4 in OP9 cells during differentiation showed that IGFBP4 upregulates adipogenesis through PPARγ, CEBPα, AGPAT2 and SREBP1 expression. We propose that this American Indian specific variant in IGFBP4 affects obesity via an increase of IGF signaling., (© 2022. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Background: The purpose of the present study was to compare the operative time for graft preparation using different techniques for graft suturing., Material and Methods: Flexor profundus tendons were harvested from fresh pig hind-leg trotters. Three different suture techniques were investigated: the Krackow stitch (K), the Whipstitch (W), and the Modified Finger-Trap suture (MFT). Tendons were sutured starting at 10 mm from the distal free end of the tendon. The suture configurations of the Krackow stitch and Whipstitch were completed with five suture throws. According to the MFT technique, the suture was wrapped five times around the tendon over a distance of 30 mm. The time required to perform a complete suture on each tendon was measured. Five independent examiners of different levels of training measured the time required for graft preparation during 3 separate occasions to determine intraobserver repeatability and interobserver reproducibility., Results: The mean time required for graft preparation following the Krackow technique was 69.1 seconds ± 18.3 SD (range 31.8-120). The Whipstitch technique took an average of 59.9 seconds ± 21.2 SD (range 27-93). The MFT suture required a mean of 29.3 seconds ± 11.4 SD for completing the suture (range 21.6-33). In all examiners the time required to complete the MFT suture was significantly less than the other suture techniques (p < 0.05). Intraobserver intraclass correlation coefficients for each examiner ranged from 0.72 to 0.83., Conclusion: Low graft preparation time is required to complete a MFT suture in a porcine tendon model. Further, time required for graft preparation using the MFT was shorter than other suturing techniques such as the Krackow and Whipstitch techniques., Clinical Relevance: The MFT suture could be used for graft set-up with the main advantage of reducing the time required in comparison with other suture techniques.

Background: Osteoarthritis (OA) is a degenerative joints disorder influenced by genetic predisposition. We reported that rs11718863 DVWA SNP was represented in Sicilian with a more severe Kellgren and Lawrence (KL) radiographic grade, displaying its predictive role as OA marker progression. Here, we describe the DVWA SNPs: rs11718863, rs7639618, rs7651842, rs7639807 and rs17040821 probably able to induce protein functional changes., Findings: Sixty-one Sicilian patients with knee OA and 100 healthy subjects were enrolled. Clinical and radiographic evaluation was performed using AKSS scores and KL. Linkage Disequilibrium (LD) analyses were performed in order to verify whether the SNPs segregate as haplotype. All DVWA SNPs'MinorAllele Frequencies (MAF) were greater than in the European. The rs7639618 SNP showed a statistical association with KL. Our analyses show that a LD exists among rs11718863 and rs7639618, as well as between rs7651842, rs7639807 and rs17040821 SNPs. We also observed that three out of the 161 individuals investigated were simultaneously homozygous carriers of the rs7651842, rs7639807 and rs17040821 MAF alleles., Conclusions: In summary, the purpose of this preliminary research was to highlight possible associations between DVWA SNPs and OA clinical and radiographic data. This work represents a multidisciplinary medicine approach to study OA where clinical, radiological and genetic evaluation could contribute to better define OA grading.