Percentage Of Collagen Research Articles

C-C motif chemokine ligand 3 is associated with atherosclerotic plaque vulnerability and increased risk of mortality

Abstract Background and Purpose Chemokines play a crucial role in the inflammatory process underlying atherosclerotic plaque formation, progression and subsequent cardiovascular events. However, the specific role of C-C motif chemokine ligand 3 (CCL3) in atherosclerosis remains largely unexplored. This study aims to explore associations between intra-plaque levels of CCL3 and plaque vulnerability as well as to investigate the prognostic role of circulating CCL3 levels in patients undergoing carotid endarterectomy. Methods The CCL3 protein expression was measured using a proximity extension assay in 207 human carotid plaques and 558 plasma samples from patients undergoing carotid endarterectomy, all enrolled in the Carotid Plaque Imaging Project (CPIP) cohort. Protein expression is quantified as normalized protein expression (NPX), an arbitrary unit on a log2-scale. Plaque components and extracellular matrix including smooth muscle cells, macrophages, neutral lipids, intraplaque hemorrhage, collagen (by histology), and matrix metalloproteinases (assessed biochemically), were quantified. Plaque vulnerability index was calculated as the sum percentage of plaque areas stained for lipids, macrophages and hemorrhage, divided by the sum percentage of smooth muscle cells and collagen. Patients were followed up to 10.2 years to record cardiovascular events and mortality. Results Plaque and circulating levels of CCL3 were higher in symptomatic patients compared to asymptomatic patients (plaque, median NPX (interquartile range, IQR): 18.2 (17.4–19.1) vs 17.2 (16.4–18.0), P<0.001; plasma, median NPX (IQR): 1.78 (1.42–2.21) vs 1.56 (1.28–1.97), P<0.001). Plaque levels of CCL3 showed positive correlations with neutral lipids, macrophages, intraplaque hemorrhage, plaque vulnerability index and matrix metalloproteinases including MMP1, MMP2, MMP9, MMP10 and TIMP1, but inversely with smooth muscle cells. Among 511 patients with complete follow-up and circulating CCL3 measures, 144 (28.2%) patients developed cardiovascular events and 123 (24.1%) patients died. Elevated circulating CCL3 levels were associated with increased risk of cardiovascular events (hazard ratio (HR) per standard deviation (SD) = 1.40, 95% CI 1.06–1.84) and mortality (HR per SD=1.77, 95% CI 1.34–2.33), respectively. Higher levels of circulating CCL3 remained independently associated with an increased risk of mortality (HR per SD=1.50, 95% CI 1.09–2.04) even after adjustment for potential confounders. Conclusions Plaque levels of CCL3 are associated with plaque vulnerability features. Circulating levels of CCL3 predict higher risk of cardiovascular events and mortality among carotid endarterectomy patients.

Effect of Photobiomodulation on an Experimental Model ofLower Limb Ischemia.

Photobiomodulation (PBM) has been shown to be promising for the promotion of angiogenesis. The present study investigated the effects of PBM on vascularization in an animal model of peripheral artery disease. Wistar rats were divided into three groups. The control group received no procedures. The ischemia group was submitted to ligation of the femoral artery of the hindleg. The ischemia + PBM group was submitted to ligation of the femoral artery followed by PBM (660 and 808 nm, 100 mW, 4 J) over the site. Animals with ischemia treated with PBM exhibited comparable results to the control group with regards to the diameter of the α-SMA+ vessels, cross-sectional area of muscle fibers, percentage of collagen and serum concentration of IL-17A, as well as similarities in terms of vertical mobility, temperature of the hindleg, number of acts of grooming, and percentage of movement, indicating a condition like that of limbs unaffected by ischemia.

Microscopic characteristics of peri- and postmortem fracture surfaces

This study investigated if microscopic surface features captured with a scanning electron microscope (SEM) effectively discriminate fracture timing. We hypothesized that microscopic fracture characteristics, including delamination, osteon pullout, and microcracks, may vary as bone elasticity decreases, elucidating perimortem and postmortem events more reliably than macroscopic analyses. Thirty-seven unembalmed, defleshed human femoral shafts from males (n=18) and females (n=2) aged 33–81 years were fractured at experimentally simulated postmortem intervals (PMIs) ranging from 1 to 60 warm weather days (250–40,600 ADH). A gravity convection oven was used to approximate tissue decomposition at 37 C and 27 C, and the resulting heat-time unit (accumulated degree hours, or ADH) was used to examine fractures in elastic/wet versus brittle/dry bone. The bones were fractured with a drop test frame using a three-point bending setup, sensors were used to calculate fracture energy, and high-speed photography documented fracture events. The following data were collected to relate fracture appearance to the biomechanical properties of bone: PMI (postmortem interval) length in ADH, temperature, humidity, collagen percentage, water loss, bone mineral density, cortical bone thickness, fracture energy, age, sex, cause of death, and microscopic fracture feature scores. SEM micrographs were collected from the primary tension zones of each fracture surface, and three microscopic fracture characteristics were scored from a region of interest in the center of the tension zone: percentage of delaminated osteons, percent osteon pullout, and number of microcracks. Multiple linear regression showed that microscopic fracture surface features are strong predictors of ADH (adjusted R-squared=0.67 for the 0 – 40,000 ADH samples; adjusted R-squared=0.92 for the 0–16,000 ADH samples). Osteon pullout is the single best predictor of ADH. Additionally, water loss is the primary driver of bone elasticity changes in low ADH samples, while collagen fibers appear to remain intact until later in the postmortem interval (approximately 40,000 ADH in this study). The results of this study indicate microscopic fracture surface analysis detects the biomechanical effects of decreased elasticity more reliably and with greater sensitivity than macroscopic analysis.

Naltrexone accelerated oral traumatic ulcer healing and downregulated TLR-4/NF-kB pathway in Wistar rats

ObjectiveTo assess the effect of naltrexone on oral mucosal healing using a traumatic ulcer model DesignWistar rats (n = 112) received distilled water (control) or naltrexone (0.5, 10, or 50 mg/kg/day). Ulcers were induced on the buccal mucosa using a round skin biopsy punch (diameter 6 mm). Euthanasia was performed on days 1, 3, 7, and 14. Healing was assessed by ulcer area, histological scores, histomorphometric analysis (number of polymorphonuclears, mononuclears, and fibroblasts), and collagen percentage. Immunohistochemistry for TLR-2, TLR-4, NF-kB, and CD31 was evaluated. Nociceptive threshold was measured daily. ResultsThe 50 mg/kg group showed reduced ulcer area on days 1 (p < 0.001), 3 (p < 0.05), and 14 (p < 0.01). In this group, there was, on day 14, an increase in the percentage of reepithelization (p = 0.043) and collagen (p < 0.05), an increase in connective tissue maturation (p = 0.016), and on day 7 an increase in fibroblasts (p < 0.001). The 10 mg/kg dose reduced the ulcer area on day 1 (p < 0.001). The 50 mg/kg group showed lower expression of TLR-4 (p < 0.001) on day 1, NF-kB on days 1 (p < 0.05) and 14 (p < 0.05), and CD31 on day 14 (p < 0.05). The 0.5 and 10 mg/kg doses reduced TLR-4 expression on day 1 (p < 0.05; p < 0.01, respectively). Nociceptive threshold increased in the 50 mg/kg group (p < 0.01). ConclusionNaltrexone enhanced traumatic oral ulcer healing by reducing TLR-4/NF-kB signaling and promoting fibroblast proliferation and collagen deposition. Additionally, naltrexone reduced pain in rats.

Gut microbiota dysbiosis links chronic apical periodontitis to liver fibrosis in nonalcoholic fatty liver disease: Insights from a mouse model.

In this study, we investigated the systemic implications of chronic apical periodontitis (CAP). CAP may contribute to the nonalcoholic fatty liver disease (NAFLD) progression through the gut microbiota and its metabolites, which are related to the degree of fibrosis. Sixteen 7-week-old male apolipoprotein E knockout (apoE-/-) mice were randomly divided into two groups: the CAP and Con groups. A CAP model was established by sealing the first- and second-maxillary molars with bacterium-containing cotton balls. Apical lesions were evaluated by micro-CT. Histological evaluations of NAFLD were performed using second harmonic generation/two-photon excitation fluorescence (SHG/TPEF) assays. Additionally, we comprehensively analyzed the gut microbiota using 16S rRNA gene sequencing and explored metabolic profiles by liquid chromatography-mass spectrometry (LC-MS). Immunofluorescence analysis was used to examine the impact of CAP on tight junction proteins and mucin expression. Transcriptome assays have elucidated gene expression alterations in liver tissues. Micro-CT scans revealed an evident periapical bone loss in the CAP group, and the total collagen percentage was increased (Con, 0.0361 ± 0.00510%, CAP, 0.0589 ± 0.00731%, p < .05). 16S rRNA sequencing revealed reduced diversity and distinct taxonomic enrichment in the CAP group. Metabolomic assessments revealed that differentially enriched metabolites, including D-galactosamine, were enriched and that 16-hydroxyhexadecanoic acid and 3-methylindole were depleted in the CAP group. Immunofluorescence analyses revealed disruptions in tight junction proteins and mucin production, indicating intestinal barrier integrity disruption. Liver transcriptome analysis revealed upregulation of Lpin-1 expression in the CAP group. This study provides comprehensive evidence of the systemic effects of CAP on liver fibrosis in NAFLD patients by elucidating alterations in the gut microbiota composition and metabolism.

Anti-inflammatory activities of phycoerythrin and phycocyanin on human fibroblast cells

Background and AimCyanobacteria are highly regarded for their nutritional value and the scientific community has shown great interest in studying their biological activity and bioactive compounds, especially pigments. They have a wide range of properties, making them potentially useful in various health applications, including dermatology. Given the growing demand for natural products and actives in the cosmetics and medical sectors, these microorganisms have emerged as a valuable source. Therefore, this study investigated the bioactivity of phycoerythrin (PE) and phycocyanin (PC) extracted from Nostoc sp. and Spirulina sp. on CCD-966SK cell lines, specifically focusing on apoptosis, necrosis, anti-inflammatory effects, and enzymatic reactions. MethodsThe study involved conducting various bioassays on a human skin fibroblast cell line (CCD-966SK) after performing cyanobacterial cell culture, pigment extraction, and fibroblast cell culture. The bioassays included the MTT test, TNF-α and IL-6 release assays, measurement of human collagen I and III, assessment of apoptosis and necrosis, and analysis of enzyme activity for superoxide dismutase (SOD) and glutathione peroxidase (GPx). ResultsThe cytotoxicity test results indicated that higher concentrations of PE and PC were associated with lower survival rates. The flow cytometry analysis confirmed a significant increase in necrotic cells and a decrease in living cells at high concentrations of both pigments. The largest percentages of collagen I for PC and PE were identified at 125 and 62.5 ppm, whereas the lowest quantities were reported at 1000 ppm for both pigments. PC and PE had the highest collagen II concentrations at 3.91 and 15.63 ppm, respectively. PC and PE had the lowest collagen II levels at 1000 ppm. PE and PC released anti-inflammatory IL-6 and TNF-a cytokines, indicating high inflammation resistance. Superoxide dismutase (SOD) and glutathione peroxidase (GPx) activity and malondialdehyde (MDA) concentrations showed that both pigments were most active at 1000 ppm. ConclusionIn conclusion, both PC and PE pigments exhibited positive effects in enhancing necrotic, anti-inflammatory, and enzymatic activity against the human skin fibroblast cell line.

Clinical and Histological Effects of Calcium Hydroxyapatite Filler in the Orofacial Region: A Study in Rats.

Calcium hydroxyapatite (CaHA) dermal filler has been increasingly used in facial aesthetic procedures. To investigate clinical and histological changes associated with calcium hydroxyapatite (CaHA) dermal filler in the orofacial region. Forty-eight female Wistar rats were divided into CaHA and control groups. The material was applied in the ventral tongue and the submandibular region; the animals were euthanized after 7, 30, and 90 days. After 7 days, yellowish nodules with a firm consistency were observed on the tongue. In 2 animals, the material migrated to the base of the tongue. Histopathological examination revealed CaHA spheres surrounded by an infiltrate, predominantly composed of macrophages. In the CaHA group, the percentage of collagen in the tongue and dermis was higher compared with the control group ( p < .05) at both 30 and 90 days. The thickness of the epidermis/dermis was also higher in the CaHA group ( p < .05). In 5 submandibular glands containing material, areas of edema and hyperemia were observed, along with infiltrates of neutrophils, lymphocytes, and plasma cells. Changes in the morphology of ducts and acini in adjacent regions were evident. CaHA exhibits satisfactory properties for filling and collagen biostimulation in the tested regions. Further studies are required to explore the potential for migration and the glandular alterations.

USP18 Curbs the Progression of Metabolic Hypertension by Suppressing JAK/STAT Pathway.

Hypertension is a pathological state of the metabolic syndrome that increases the risk of cardiovascular disease. Managing hypertension is challenging, and we aimed to identify the pathogenic factors and discern therapeutic targets for metabolic hypertension (MHR). An MHR rat model was established with the combined treatment of a high-sugar, high-fat diet and ethanol. Histopathological observations were performed using hematoxylin-eosin and Sirius Red staining. Transcriptome sequencing was performed to screen differentially expressed genes. The role of ubiquitin-specific protease 18 (USP18) in the proliferation, apoptosis, and oxidative stress of HUVECs was explored using Cell Counting Kit-8, flow cytometry, and enzyme-linked immunosorbent assays. Moreover, USP18 downstream signaling pathways in MHR were screened, and the effects of USP18 on these signaling pathways were investigated by western blotting. In the MHR model, total cholesterol and low-density lipoprotein levels increased, while high-density lipoprotein levels decreased. Moreover, high vessel thickness and percentage of collagen were noted along with increased malondialdehyde, decreased superoxide dismutase and catalase levels. The staining results showed that the MHR model exhibited an irregular aortic intima and disordered smooth muscle cells. There were 78 differentially expressed genes in the MHR model, and seven hub genes, including USP18, were identified. USP18 overexpression facilitated proliferation and reduced apoptosis and oxidative stress in HUVECs treated with Ang in vitro. In addition, the JAK/STAT pathway was identified as a USP18 downstream signaling pathway, and USP18 overexpression inhibited the expression of JAK/STAT pathway-related proteins. Conclusively, USP18 restrained MHR progression by promoting cell proliferation, reversing apoptosis and oxidative stress, and suppressing the JAK/STAT pathway.

Effect of poly-L-lactic acid and polydioxanone biostimulators on type I and III collagen biosynthesis.

Safe, effective, and biocompatible minimally invasive procedures with the potential to stimulate collagen production have been made to recover dermal thickness and skin quality. The main of this animal model experiment was to observe the effect of poly-L-lactic acid (PLLA) and polydioxanone (PDO) biostimulators in collagen I and III after hypodermal injection. Sixteen adult female rats (Wistar) were randomized into four groups and had dorsal treatment with: G1: hypodermic subcision (HS) only; G2: HS and PLLA hypodermic injection (HI), G3: HS and PDO HI; G4: Control, with no treatment. In histochemical, it was observed hypodermal and dermal tissue in more organized thickness in G3 and in G4 when compared to G1 and G2. There was few difference in G1 compared to G4. The tissue of G2 showed irregularities in the arrangement of collagen fibers, less defined structure and lower distribution of type I collagen compared to the other groups. There is a greater tendency for the proportions of type III collagen among tissues treated with both biostimulators (G2 and G3). PLLA and PDO had relatively similar percentages of collagen when compared to G4. The amount of type I collagen was higher in tissues treated with subcision, while type III collagen was higher in tissues treated with both biostimulators. G3 showed better performance in collagen production, although small, when compared with G2.

Abstract 6180: Tumor microenvironment phenotypes predict recurrence in African Americans diagnosed with colorectal cancer: A comparative study of manual grading with digital pathology

Abstract African Americans face a higher rate of colorectal cancer (CRC) incidence and mortality compared to other ethnicities. Identifying key prognostic factors is essential for better outcomes in this group. This study focused on the prognostic value of stromal grading in the tumor microenvironment (TME) of African American CRC patients, using both manual grading and digital pathology. A retrospective analysis of CRC cases in African Americans was conducted. Two cohorts were established: a control cohort (patients without recurrence) and a recurrence cohort (patients with recurrence). Stromal grading was performed manually, categorizing tumors into Grade 1 (low risk), Grade 2 (moderate risk), and Grade 3 (high risk), and we also quantified the TME percentages by digital pathology: myxoid, collagen, immune and tumor. In the control cohort (N=40), stromal grading identified 30 Grade 1, 7 Grade 2, and 3 Grade 3 cases. Conversely, the recurrence cohort (N=17) demonstrated a starkly different distribution, with 1 Grade 1, 3 Grade 2, and 13 Grade 3 cases. The Chi-square statistic was notably significant (p&amp;lt;0.0001), indicating a strong association between higher stromal grades and the likelihood of recurrence. By unpaired t-test, high myxoid stroma (p&amp;lt;0.0001), low collagen (p&amp;lt;0.0001), and low tumor (p=0.0002) by digital pathology were associated with recurrence. TME phenotypes are a significant prognostic and recurrence indicator in African American patients with CRC. The marked difference in stromal grades and digital pathology features between the control and recurrence cohorts suggests that higher stromal grades, along with higher myxoid, lower collagen and lower tumor percentages are associated with an increased risk of recurrence. These findings underscore the need for tailored therapeutic strategies and more vigilant surveillance in African American patients exhibiting high-risk TME phenotypes. The efficacy of manual and digital pathology evaluation of the TME also warrants further exploration to enhance prognostic accuracy and treatment personalization. Citation Format: Dongling Wu, Rong Xia, Mouyed Alawad, Lili Lee, Tamas Gonda, Wenqing Cao, Sean Hacking. Tumor microenvironment phenotypes predict recurrence in African Americans diagnosed with colorectal cancer: A comparative study of manual grading with digital pathology [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6180.

The role of preoperative MRI in endoscopic transnasal transsphenoidal hypophysectomy of pituitary adenoma.

The trans-sphenoidal approach, commonly used for removing pituitary adenomas, has become a widely accepted and successful method. In recent years, the endoscopic trans-sphenoidal technique has emerged as a minimally invasive surgical approach for pituitary adenoma removal. The majority of pituitary adenomas exhibit a soft consistency and can be successfully extracted with aspiration and curettage using the trans-sphenoidal approach. However, a subset of around 5-15% of these adenomas possess a solid and fibrous texture. The occurrence of firm and fibrous adenomas is relatively common; unfortunately, there are no reliable predictors to identify them preoperatively. The ability to forecast the reliability of magnetic resonance imaging (MRI) holds promise for improving prior preparation and impacts the extent of resection. A cross-sectional analysis of the investigation of magnetic resonance imaging (MRI) in relation to cancer histology was performed on 68 patients who had endoscopic trans-nasal excision for nonfunctional adenomas. The determination of an intensity ratio was performed by employing quantitative estimates of MRI signal intensity obtained from both the adenoma and pons. During the surgical procedure, a series of sequential-graded procedures were used for the removal of tumours with varying consistencies. Softer tumours were addressed using the Suction technique (R1), while tumours of intermediate consistency were treated using curettes (R2). In order to evaluate the fibrotic content of firmer tumours, the utilization of Cavitron Ultrasound Surgical Aspirator(CUSA), and/or other micro-instruments (R3) was employed, with the histologic collagen fraction being quantified. In order to investigate and analyse the data, a statistical analysis was conducted. A predictive relationship between resection category and both intensity ratio, and collagen percentage was noted. The primary objective of this study was to determine the appropriate cutoff criteria for clinical utilization, as well as to investigate the association between intensity ratios and collagen percentage. Tumors with ratios ≤ 1.6 on the T2-weighted image and collagen content > 5.3% required more meticulous and sharp dissection for resection. The utilization of MRI analysis may offer some assistance, but not conclusive, in the prediction of tumour consistency.

Evaluation of the use of chamomile in isolation and in association with laser photobiomodulation in the healing of rats oral mucosa.

Laser photobiomodulation (LPBM) has been shown to be one of the possible modulating agents of inflammation. Similarly, medicinal plants, such as chamomile (Matricaria recutita) are also used with the same purpose. To evaluate tissue repair in the dorsum of the tongue of rats under topical use of chamomile alone and in association with LPBM. Seventy-five male Wistar rats received a standardized wound on the dorsum of the tongue and were allocated into experimental groups: Control (G1), Chamomile Fluid extract (G2), Chamomile Infusion (G3), Laser (G4), Chamomile Infusion + Laser (G5). Euthanasia was done on days 3, 7, and 14 after surgery. Ulcers were evaluated and measured with a caliper. Sections stained with hematoxylin and eosin and Picrosirius Red allowed evaluation of edema, inflammatory infiltrate, cellularity, and re-epithelialization and characterization of total collagen. Histomorphometric analysis of the percentage of total collagen, the distance from the basal layer to the epithelial surface, and the thickness of the stratum corneum were performed. The G2 and G4 groups modulated the exudative and proliferative phases of inflammation, both clinically and histologically. The G3 and G5 groups did not show significant differences in relation to the G1 group in most of the evaluated parameters. Chamomile fluid extract and LPBM alone showed better clinical and histological responses for tissue repair than the association between these therapeutic modalities. There were differences in the parameters of clinical, histological, and histomorphometric patterns between the experimental groups of the present investigation. The LPBM proved to be superior in the performed analysis.

Morphological and ultrastructural investigation of the posterior atlanto-occipital membrane: Comparing children with Chiari malformation type I and controls.

The fibrous posterior atlanto-occipital membrane (PAOM) at the craniocervical junction is typically removed during decompression surgery for Chiari malformation type I (CM-I); however, its importance and ultrastructural architecture have not been investigated in children. We hypothesized that there are structural differences in the PAOM of patients with CM-I and those without. In this prospective study, blinded pathological analysis was performed on PAOM specimens from children who had surgery for CM-I and children who had surgery for posterior fossa tumors (controls). Clinical and radiographic data were collected. Statistical analysis included comparisons between the CM-I and control cohorts and correlations with imaging measures. A total of 35 children (mean age at surgery 10.7 years; 94.3% white) with viable specimens for evaluation were enrolled: 24 with CM-I and 11 controls. There were no statistical demographic differences between the two cohorts. Four children had a family history of CM-I and five had a syndromic condition. The cohorts had similar measurements of tonsillar descent, syringomyelia, basion to C2, and condylar-to-C2 vertical axis (all p>0.05). The clival-axial angle was lower in patients with CM-I (138.1 vs. 149.3 degrees, p = 0.016). Morphologically, the PAOM demonstrated statistically higher proportions of disorganized architecture in patients with CM-I (75.0% vs. 36.4%, p = 0.012). There were no differences in PAOM fat, elastin, or collagen percentages overall and no differences in imaging or ultrastructural findings between male and female patients. Posterior fossa volume was lower in children with CM-I (163,234 mm3 vs. 218,305 mm3, p<0.001), a difference that persisted after normalizing for patient height (129.9 vs. 160.9, p = 0.028). In patients with CM-I, the PAOM demonstrates disorganized architecture compared with that of control patients. This likely represents an anatomic adaptation in the presence of CM-I rather than a pathologic contribution.

Characterization of an in vitro engineered ligament model

In vivo tendon and ligament research can be limited by the difficultly of obtaining tissue samples that can be biochemically analyzed. In this study, we characterize the most widely used in vitro engineered ligament model. Despite previous works suggesting multiple passages change gene expression in 2D primary tenocytes, we found no relationship between passage number and expression of classical tendon fibroblast markers across different biological donors. When engineered into 3D ligaments, there was an increase in maximal tensile load between 7 and 14 days in culture, that corresponded with an increase in collagen content. By contrast, percent collagen increased logarithmically from Day 7 to Day 14, and this was similar to the increase in the modulus of the tissue. Importantly, there was no relationship between passage number and mechanical function or collagen content in the two independent donors tested. These results suggest that the model develops quickly and is reliable across differing passage numbers. This provides the field with the ability to 1) consistently determine functional changes of interventions out to passage number 10; and 2) to time interventions to the appropriate developmental stage: developing/regenerating (Day 7) or mature (Day 14) tissue.

Melatonin acts through different mechanisms to control oxidative stress and primordial follicle activation and survival during in vitro culture of bovine ovarian tissue

This study aims to evaluate the effects of melatonin and its mechanisms of action on preantral follicle activation and survival, stromal cell density and collagen distribution in extracellular matrix (ECM). The involvement of melatonin receptors and mTORC1 pathway in these procedures were also investigated. To this end, ovarian fragments were cultured for six days in α‐MEM+ alone or supplemented with 1000 pM melatonin, 1000 pM melatonin with 1000 pM luzindole (inhibitor of melatonin receptors), or 1000 pM melatonin with 0.16 µg/ml rapamycin (mTORC1 inhibitor). At the end of culture period, tissues were processed for classical histology, and the follicles were classified as normal or degenerated, as well as in primordial or growing follicles. The ovarian stromal cell density and ECM collagen distribution were also evaluated. Samples of ovarian tissues were also destined to measure the levels of thiol and mRNA for CAT, SOD, GPX1 and PRDX1, as well as the activity of antioxidant enzymes CAT, SOD, and GPX1. The results demonstrated that ovarian tissues cultured with melatonin, melatonin with luzindole or melatonin with rapamycin had significantly higher percentage of morphologically normal follicles than those cultured in control medium (α‐MEM+). However, the presence of either luzindole or rapamycin, did not block the positive effects of melatonin on follicle survival (P > 0.05). Although the presence of melatonin in culture medium reduced the percentage of primordial follicles and increased the percentage of development follicles, these positive effects of melatonin were blocked by either luzindole or rapamycin (P < 0.05). Melatonin, melatonin with luzindole or melatonin with rapamycin did not influence the number of ovarian stromal cells. In contrast, melatonin significantly increased the percentages of collagen in ovarian tissues, but the positive effects of melatonin were blocked by either luzindole or rapamycin. Tissues cultured with melatonin and rapamycin had higher levels of mRNA for CAT and lower GPx activity when compared to those cultured in control medium. In conclusion, melatonin promotes primordial follicle activation, increases collagen fiber in ECM of in vitro cultured bovine ovarian tissue through its membrane-coupled receptors and mTORC1. Oppositely, melatonin increase follicles survival by acting through other pathways, since it can pass through cell membranes and directly regulate oxidative stress.

Chronic caffeine intake alters collagen deposition in long bones: an experimental preliminary study

This study explores the impact of chronic caffeine intake on collagen deposition in long bones and correlates it with morphometric and metabolic alterations in Wistar rats. Animals had either regular drinking water (Control group) or caffeinated drinking water in a 300 mg/L dose (Caffeine group) for 12 wk. Using Picro-Sirius red collagen staining, cortical (78.07 ± 1.27 vs 82.12 ± 1.22) and trabecular (74.29 ± 1.09 vs 77.97 ± 1.03) collagen percentages in the caffeine group were markedly (p = 0.0448 and p = 0.0351; respectively) lower than the control group. Caffeine intake induced apparent alterations in the microstructure of cortical diaphysis and femoral head trabecular density as determined by micro-CT. In caffeinated animals, levels of bone-related metabolic biomarkers including BALP (1.41 ± 0.06 vs 1.12 ± 0.07), CTX (313.5 ± 20.78 vs 224.3 ± 27.19), DPD (11.61 ± 0.72 vs 9.26 ± 0.62), BGP (10.35 ± 0.43 vs 9.14 ± 0.34), and TRAP (4.34 ± 0.08 vs 4.01 ± 0.11) were increased (p = 0.0136, p = 0.0262, p = 0.0343, p = 0.0571, and p = 0.0461; respectively) as compared to control animals. Except for BALP (p = 0.0175) and DPD (p = 0.0423), the Pearson correlation between collagen bone deposition and microstructures and metabolic changes was not statistically significant. Collectively, chronic caffeine consumption restricts cortical and trabecular collagen deposition in long bones leading to microstructure and metabolic alterations.

Dapagliflozin treatment and cardiovascular outcome in RBP4/TTRVal30Met (transthyretin cardiac amyloidosis) mice.

Whether sodium-glucose co-transporter 2 inhibitors are effective for heart failure caused by ATTR-CA (transthyretin cardiac amyloidosis) remains uncertain. The aim of this study is to investigate the cardiovascular prognosis in ATTR-CA mice model with dapagliflozin treatment. Humanized RBP4/TTRVal50Met and RBP4/TTR mice models were constructed with clustered regularly interspaced short palindromic repeats and associated Cas9 endonuclease (CRISPR-Cas9) techniques and multiple generations breeding. A total of 6 RBP4/TTR mice received placebo treatment, when 12 RBP4/TTRVal50Met received dapagliflozin (1mg/kg/day, 6 mice) and placebo (6 mice) treatment. Fasting glucose, intraperitoneal glucose tolerance test, and plasma brain natriuretic peptide (BNP) concentration were measured at Day 0, Week 2, and Week 4. BNP, transforming growth factor-beta (TGF-β), collagen type I alpha 1 (COL1A1) protein levels, and Cola1, TGFβ1, TNFα, IL-1β, BNP relative quantities in cardiac, along with cardiac pathology examination including right ventricular collagen percentage, ventricular septum thickness, left ventricular wall thickness, and left ventricular internal diameter were measured at Week 4 after treatment procedure. All 18 mice completed the experiment. The baseline characteristics were balanced among three treatment groups. In placebo-treated mice, the cardiac BNP relative quantity was significantly higher in RBP4/TTRVal50Met mice than RBP4/TTR mice (RBP4[KI/KI], TTR [KI/KI]: 0.72±0.46, RBP4[KI/KI], TTRVal50Met [KI/KI]: 1.44±0.60, P=0.043), indicating more significant heart failure progression in ATTR-CA mice than normal mice. In ATTR-CA mice, the cardiovascular prognosis measurements including heart failure (plasma BNP concentration and relative quantities of BNP), cardiac inflammation (relative quantities of Cola1, TGFβ1, TNFα, and IL-1β), and pathological changes (right ventricular collagen percentage, ventricular septum thickness, left ventricular wall thickness, and left ventricular internal diameter) were statistically comparable between those under dapagliflozin and placebo treatment. Dapagliflozin did not improve cardiovascular prognosis including the progression of heart failure, cardiac inflammation, and pathological changes in ATTR-CA mice compared with placebo. The results of this study were not in support of dapagliflozin's therapeutic effects for ATTR-CA. More pre-clinical and clinical researches to validate these findings and demonstrate the underlying mechanisms are still required.

Indoxyl sulfate induced frailty in patients with end-stage renal disease by disrupting the PGC-1α-FNDC5 axis.

Sarcopenia or frailty is common among patients with chronic kidney disease (CKD). The protein-bound uremic toxin indoxyl sulfate (IS) is associated with frailty. IS induces apoptosis and disruption of mitochondrial activity in skeletal muscle. However, the association of IS with anabolic myokines such as irisin in patients with CKD or end-stage renal disease (ESRD) is unclear. This study aims to elucidate whether IS induces frailty by dysregulating irisin in patients with CKD. The handgrip strength of 53 patients, including 28 patients with ESRD, was examined. Serum concentrations of IS and irisin were analyzed. CKD was established in BALB/c mice through 5/6 nephrectomy. Pathologic analysis of skeletal muscle was assessed through haematoxylin and eosin and Masson's trichrome staining. Expression of peroxisome proliferator-activated receptor-gamma coactivator PGC-1α and irisin were analyzed using real-time polymerase chain reaction and Western blotting. Handgrip strength was lower among patients with ESRD than among those without ESRD. In total, 64.3% and 24% of the patients in the ESRD and control groups had low handgrip strength, respectively (p < 0.05). Serum concentrations of IS were significantly higher in the ESRD group than in the control group (222.81 ± 90.67 μM and 23.19 ± 33.28 μM, respectively, p < 0.05). Concentrations of irisin were lower in the ESRD group than in the control group (64.62 ± 32.64 pg/mL vs. 99.77 ± 93.29 pg/mL, respectively, p < 0.05). ROC curves for low handgrip strength by irisin and IS were 0.298 (95% confidence interval (CI): 0.139-0.457, p < 0.05) and 0.733 (95% CI: 0.575-0.890, p < 0.05), respectively. The percentage of collagen was significantly higher in mice with 5/6 nephrectomy than in the control group. After resveratrol (RSV) treatment, the percentage of collagen significantly decreased. RSV modulates TGF-β signaling. In vitro analysis revealed that IS treatment suppressed expression of PGC-1α and FNDC5 in a dose-dependent manner, whereas RSV treatment attenuated IS-induced phenomena in C2C12 cells. IS was positively correlated with frailty in patients with ESRD through the modulation of the PGC-1α-FNDC5 axis. RSV may be a potential drug for reversing IS-induced suppression of the PGC-1α-FNDC5 axis in skeletal muscle.

Characteristics of intramuscular collagen in calf-fed Nellore bulls and steers throughout the finishing phase

This study evaluated collagen solubility and gene expression of biomarkers for intramuscular collagen (IMCT) deposition and remodeling in the Longissimus muscle of bulls and steers through the finishing phase. Thirty-six Nellore calves were used (18 bulls and 18 steers), and six of each sexual condition were randomly assigned to be harvested at 0, 100, or 200 days on feed (DOF) to evaluate collagen characteristics in different time points throughout the finishing phase. Bulls showed a greater collagen solubility than steers (P = 0.03). The gene expression of fibrogenic markers (TGFβ1, COL1A1, and COL3A1) and IMCT remodeling mediators (MMPII, TIMPII, and LOX) were not affected by sexual condition or DOF (P > 0.05). Our data indicate that young Nellore bulls have a higher percentage of soluble intramuscular collagen, possibly due to higher collagen remodeling associated with a faster growth rate and muscle hypertrophy. Moreover, castration and DOF did not modify mRNA levels of fibrogenic and collagen remodeling markers.

Effect of Different Roughage Sources in Fermented Total Mixed Ration and Energy Intake on Meat Quality, Collagen Solubility, Troponin T Degradation, and Fatty Acids of Native Thai Cattle Longissimus Muscle.

The effects of roughage sources in the fermented total mixed ration (FTMR) and the level of energy intake on meat quality, collagen solubility, and troponin T degradation in longissimus thoracis (LT) muscle of native Thai cattle (NTC) were investigated. Results showed that roughage source affected fatty acid composition in the LT muscle (p < 0.05), as NTC fed Pakchong 1-Napier-based FTMR had higher monounsaturated fatty acid content and ω 6:ω 3 ratio. The high-energy ad libitum group had lower drip loss, lower shear force, and higher percent collagen solubility (p < 0.05). However, energy intake had no effect on troponin T degradation and fatty acid composition (p > 0.05). Longer aging of 14 days showed lower shear force values, higher collagen solubility, and troponin T degradation rate but higher cooking loss (p < 0.01). In conclusion, the meat quality of NTC could be improved by ad libitum feeding with NG-FTMR, as their meat had higher MUFA content, lower drip loss, lower shear force, and higher collagen solubility. In addition, the tenderness of NTC meat could be further improved by longer aging of 14 days post-mortem.