Collagenous Connective Tissue Research Articles

Unveiling putative modulators of mutable collagenous tissue in the brittle star Ophiomastix wendtii: an RNA-Seq analysis

Collagenous connective tissue, found throughout the bodies of metazoans, plays a crucial role in maintaining structural integrity. This versatile tissue has the potential for numerous biomedical applications, including the development of innovative collagen-based biomaterials. Inspiration for such advancements can be drawn from echinoderms, a group of marine invertebrates that includes sea stars, sea cucumbers, brittle stars, sea urchins, and sea lilies. Through their nervous system, these organisms can reversibly control the pliability of their connective tissue components (i.e., tendons and ligaments) that are composed of mutable collagenous tissue (MCT). The variable tensile properties of the MCT allow echinoderms to perform unique functions, including postural maintenance, reduction of muscular energy use, autotomy to avoid predators, and asexual reproduction through fission. The changes in the tensile strength of MCT structures are specifically controlled by specialized neurosecretory cells called juxtaligamental cells. These cells release substances that either soften or stiffen the MCT. So far, only a few of these substances have been purified and characterized, and the genetic underpinning of MCT biology remains unknown. Therefore, we have conducted this research to identify MCT-related genes in echinoderms as a first step towards a better understanding of the MCT molecular control mechanisms. Our ultimate goal is to unlock new biomaterial applications based on this knowledge. In this project, we used RNA-Seq to identify and annotate differentially expressed genes in the MCT structures of the brittle star Ophiomastix wendtii. As a result, we present a list of 16 putative MCT modulator genes, which will be validated and characterized in forthcoming functional analyses.

Wound healing activity of Ipomoea staphylina leaves extract in Wistar rats

Wound repair is the practice of repairing the skin and other soft tissues after an injury. An inflammatory response is activated after an injury, and cells below the dermis (deepest skin layer) begin to produce more collagen (connective tissue). A scientific assessment was made for the wound-healing potential of Ipomoea staphylina leaves extract, using ethanol. The crude extract was made into a 2.5% and 5% (w/w) ointment and tested for wound healing activity in Wistar rats using excision and incision wound models. In the excision wound model, the period of epithelialization was reduced and an increase in wound contraction rate was observed in the extract treated groups (III and IV). On the tenth day, tissue from the excision wound area was taken and processed for the assessment of pro-inflammatory cytokines (TNF-α), and it was found that there was a notable decrease in the TNF-α concentration in the extract-treated animals. In an incision wound model, the breaking strength was significantly increased in animals treated with 2.5% and 5% (w/w) ointment of I. staphylina leaves extract. The extract significantly promoted fibroblast migration in an in vitro experiment (scratch assay), which may have been caused by the presence of flavonoids.

137 Possible mechanisms underlying enhanced intramuscular adipogenesis in Wagyu cattle

Abstract The quality of beef hinges significantly on the abundance of intramuscular fat and tenderness. Intramuscular fat accumulation results directly from intramuscular adipogenesis, while tenderness is adversely affected by collagenous intramuscular connective tissue. Both intramuscular fat and connective tissue originate from fibro/adipogenic progenitors (FAPs), a group of bipotent progenitors. Wagyu cattle are renowned for their tender beef with rich intramuscular fat, likely owing to the distinctive features of Wagyu FAPs. Our study has identified fewer myogenic cells but more FAPs with greater adipogenic potential in Wagyu compared with Angus muscle. Furthermore, single-cell transcriptomic analysis identified that Wagyu FAPs exhibit greater adipogenic and decreased fibrogenic characteristics compared with FAPs in Brahman cattle, which generally produce low-quality beef with limited intramuscular fat and tenderness. Interestingly, Wagyu muscle also contains more abundant vascular cells, crucial for supporting adipogenesis. Mechanistic studies have unveiled several signaling pathways, including the bone morphogenetic protein signaling and the alternative complement pathway of the complement system, which likely contributes to the heightened adipogenic programming of Wagyu FAPs and is subject to epigenetic regulation.

Growth factor-enhanced matrix combined either with free connective tissue graft or with collagen membrane in the treatment of Miller class II gingival recession

Abstract Purpose The goal of the study was to examine the efficacy of collagen membrane or free connective tissue graft in conjunction with growth factor-enhanced matrix (GEM) for treating Miller class two gingival recession. Patients and methods Twenty sites in 10 patients, with class II Miller gingival recession were allocated randomly to be treated by (GEM21s) with free connective tissue graft (group I) or GEM21s with collagen membrane (group II). Recession width, recession depth, height of keratinized tissue and digital measurements were recorded at baseline, 1, 3, and 6 months. Probing pocket depth, clinical attachment level, gingival thickness, and radiographic parameters were recorded at baseline and 6 months. Root coverage percentage were recorded 6 months postoperative and wound healing index were recorded at 1, 3, and 6 months postoperative. Results Both groups showed significant improvement in all clinical parameters and increase in bone level was observed after 6 months compared to baseline measures. There was no statistically significant difference between both groups except for recession depth at 1 and 3 months after surgery and height of keratinized tissue at 1, 3, and 6 months in favor of group I. Radiographically there was a statistically significant difference (P ≤ 0.05) in amount of labial bone gain and bone density in favor of group I compared to group II. Conclusion GEM21s combined either with free connective tissue graft or collagen membrane led to favorable gingival coverage and radiographical results with better results for free connective tissue graft with GEM21s.

Volumetric assessment of volume stable collagen matrix in maxillary single implant site development: A randomized controlled clinical trial.

The stability of soft tissue volume around dental implants is an important factor for the final esthetic outcome. The main objective of this study was to compare volume stable collagen matrix (VCMX) versus connective tissue graft (CTG) in the augmentation of soft tissue profiles in single implant sites with a class I Siebert ridge defect. Twenty patients (14 females and 6 males) were enrolled in the present study. After implant placement and augmentation of the buccal defect by VCMX or CTG, post-operative evaluation of the volumetric changes at the augmented implant site was carried out at 3, 6, and 9 months as primary outcome, clinical and radiographic soft tissue thickness were carried out at baseline and 9-month intervals, visual analog scale (VAS) and oral health impact profile-14 (OHIP14) were recorded 2 weeks after the surgery. A statistically significant difference in soft tissue volume was found between baseline and 3, 6, and 9 months postoperatively in both groups with the highest value at 9 months (136.33 ± 86.80) (mm3) in VCMX and (186.38 ± 57.52) (mm3) in CTG. Soft tissue thickness was significantly increased in both groups at 9 months in comparison to baseline. However, there was a significantly higher increase in soft tissue thickness at 9 months in CTG (3.87 ± 0.91) than in VCMX (2.94 ± 0.31). Regarding the radiographic soft tissue thickness, there was a statistically significant increase in both groups at 9 months in comparison to baseline. However, there was a statistically higher increase in the radiographic soft tissue thickness at 9 months in CTG (3.08 ± 0.97) than in VCMX (2.37 ± 0.29). VAS showed a statistically lower value in VCMX (0.4 ± 0.7) than CTG (2.8 ± 1.48). The OHIP recorded lower values in the VCMX group than the CTG group with no statistical significance. In addition, there was no difference in the PES between the two groups. The present study showed that CTG and VCMX were both effective in soft tissue augmentation around implants in the esthetic zone. However, CTG proved more efficient in increasing peri-implant soft tissue volume and mucosal thickness around single implants at a 9-month follow-up period. VCMX was associated with less pain or discomfort and reduced patient morbidity, as reflected by the significantly reduced VAS value in the VCMX group.

Microstructural evolution and mechanics behavior of postmortem meat subjected to resonance

The tenderness of meat is determined by muscle components, mainly including myofibrillar and connective tissues. In the present study, the microstructures evolution and mechanics behavior of yak postmortem muscle subjected to resonance vibration treatment was investigated by experimental and numerical investigation, in order to illustrate the resonance tenderization mechanism and the mechanical relationship of muscle components. The results show that there are three orders of natural frequencies of the raw yak meat, and the resonance frequency is determined as 26 Hz. By vibration treatment at this frequency, the shear force and MFI of the vibrated meat is 35.70 N and 62, respectively, exhibiting remarkable tenderizing effect compared with the raw sample and other frequencies treated samples. The microstructures and vibration modal simulation have revealed that the stress concentration happens in the junction of muscle fibers and extracellular matrix (collagenous connective tissues), leading to the detachment of endomysium connective tissues from muscle fibers and the disappearance of M-bands in myofibrils, thus resulting the significant tenderization. The resonance also causes the stress-strain behaviors of meat transiting from plastic deformation to elastic deformation. The experimental and numerical results reveal that the physical destruction of the cross-linkings or interactions between muscle fibers and endomysium connective tissues induced by resonance, plays a crucial role in the detachment of muscle components, transformation of stress-strain behavior and the significant tenderization effect.

Correlation between the region of interest in digital radiography, Hounsfield units, and histological maturation on Wistar rats submitted to tibial fracture

Objective: Study the relationship between the region of interest (ROI) and Hounsfield units (HU) in the tibial bone callus. Methods: Twenty-one adult Wistar rats were submitted to tibial fracture. The fracture was radiographed after their euthanasia, and the bone calluses were analyzed histologically after being stained with hematoxylin and eosin. Euthanasia occurred between the 2nd and 6th weeks after fracture and fixation, thus obtaining various consolidation stages. Histologically, vessels, chondroblasts, connective tissue (collagen), maximum size of the chondrocyte, and the concentration of chondrocytes were quantified. Results: It was observed that the higher the HU, the more mature and closer to bone consolidation it is, proving that the use of ROI and bone callus measurement with HU is reliable for the histological process of maturation of the bone callus and can be safely used as proof of evolution of the bone healing process. Conclusion: The ROI was successfully used in digital radiography to observe HU in fractured bones. Level of Evidence IV; Therapeutic Studies; Case Series.

Dynamic analyses of a soft tissue-implant interface: Biological responses to immediate versus delayed dental implants.

To qualitatively and quantitatively evaluate the formation and maturation of peri-implant soft tissues around 'immediate' and 'delayed' implants. Miniaturized titanium implants were placed in either maxillary first molar (mxM1) fresh extraction sockets or healed mxM1 sites in mice. Peri-implant soft tissues were evaluated at multiple timepoints to assess the molecular mechanisms of attachment and the efficacy of the soft tissue as a barrier. A healthy junctional epithelium (JE) served as positive control. No differences were observed in the rate of soft-tissue integration of immediate versus delayed implants; however, overall, mucosal integration took at least twice as long as osseointegration in this model. Qualitative assessment of Vimentin expression over the time course of soft-tissue integration indicated an initially disorganized peri-implant connective tissue envelope that gradually matured with time. Quantitative analyses showed significantly less total collagen in peri-implant connective tissues compared to connective tissue around teeth around implants. Quantitative analyses also showed a gradual increase in expression of hemidesmosomal attachment proteins in the peri-implant epithelium (PIE), which was accompanied by a significant inflammatory marker reduction. Within the timeframe examined, quantitative analyses showed that connective tissue maturation never reached that observed around teeth. Hemidesmosomal attachment protein expression levels were also significantly reduced compared to those in an intact JE, although quantitative analyses indicated that macrophage density in the peri-implant environment was reduced over time, suggesting an improvement in PIE barrier functions. Perhaps most unexpectedly, maturation of the peri-implant soft tissues was a significantly slower process than osseointegration.

Results of surgical treatment of hernias of the anterior abdominal wall in patients with connective tissue dysplasia syndrome

Relevance. The problem of surgical treatment of hernias of the anterior abdominal wall remains relevant. Despite the introduction of new technologies, the results of surgical interventions cannot be considered satisfactory. The development of hernias and their relapses is facilitated by connective tissue dysplasia syndrome, which manifests as phenotypic signs, disorders of the autonomic nervous system, and morphological changes in the structure of the connective tissue.Objective. To improve the results of surgical treatment of hernias of the anterior abdominal wall in patients with connective tissue dysplasia syndrome.Materials and methods. The study was based on clinical observations and studies of 497 patients with hernias of the anterior abdominal wall. Inguinal (29 %) and postoperative hernias (21 %) predominated. In the structure of recurrent hernias, postoperative (47 %) and inguinal (42 %) hernias were more often observed. Hernioautoplasty was performed in 31 %, plastic surgery using a mesh endoprosthesis — in 54 %, endoscopic hernioplasty — in 15 % of patients. The severity of the clinical, morphological, and metabolic signs of connective tissue dysplasia syndrome was assessed. The concentration of free hydroxyproline in the blood was determined as a marker of connective tissue metabolism. The state of the autonomic nervous system was assessed using variational pulsometry. To assess the state of collagen in connective tissue, a morphological study of the aponeuroses of the muscles of the anterior abdominal wall was performed in the clinic and experiment.Results. The overall incidence of dysplasia in hernias of the anterior abdominal wall was 48 %. The highest incidence of connective tissue dysplasia was detected in recurrent hernias (70 %). Studies of the level of free hydroxyproline in abdominal hernias have shown the dependence of this marker on the degree of connective tissue dysplasia. In cases of recurrent hernias, the highest intensity of collagen biodegradation was observed. A study of the functional state of the autonomic nervous system revealed a predominance of sympathetic activity in abdominal wall hernias with an increase in the tension index by more than 4 times. An increase in the severity of dysplasia is accompanied by an increase in sympathetic activity. Histological studies in patients with abdominal hernias revealed characteristic changes in the morphological structure of the connective tissue. The average thickness, specific area, brightness of staining, and orientation of collagen fibers in postoperative and recurrent hernias were significantly different compared to the group with primary hernias (p˂0.05). Additionally, an experiment was performed on 30 laboratory animals (white rats) to determine the optimal conditions for healing of the aponeurosis of the anterior abdominal wall. The activating effect of prozerin on the process of histogenesis in the wound was revealed. Based on the results of the study, the surgical treatment of hernias was assessed depending on the type of hernioplasty and the severity of connective tissue dysplasia. It has been established that with increasing severity of dysplasia, the frequency of relapses increases by 2.2–3.4 times. With the progression of dysplasia, the frequency of relapses increased by more than twice. The use of the endoscopic hernioplasty method reduced the risk of recurrent hernias by 9 times.Conclusion. In externally localized hernias, the incidence of connective tissue dysplasia ranges from 34 % to 70 %. In patients with connective tissue dysplasia syndrome after hernia autoplasty, the frequency of hernia recurrence is 3.4 times higher. Drug correction of sympathetic activity along with alloplasty increased collagen synthesis intensity by 36 %. The choice of hernia repair method should be determined by the type of hernia, the age of the patient, and the severity of connective tissue dysplasia.

Analysis of the treatment of RT2 recessions with a xenogeneic collagen matrix vs. connective tissue graft combined with a coronally advanced flap. A double-blinded randomized clinical trial

ObjectivesTo compare the clinical efficacy in terms of mean root coverage in RT2 recession treated with a coronally advanced flap combined with a xenogeneic collagen matrix versus a connective tissue graft.Materials and methodsA total of 20 patients were randomized to receive one of two treatments: coronally advanced flap + xenogeneic collagen matrix (test group) and coronally advanced flap + connective tissue graft (control group). Patient-related outcomes measures and professional aesthetic assessment by root esthetic score were performed. A descriptive and analytical statistical analysis of the variables was performed.ResultsAt 12 months, the mean root coverage was 56.48% in the test group and 69.72% in the control group (p = 0.048), with a 35% and 40% complete root coverage in the xenogeneic collagen matrix and connective tissue graft, respectively. Test group presented less pain (3.65 vs. 5.2 VAS units) (p = 0.015) and less surgical time (45 vs. 49.15 min) (p = 0.004) than control group.ConclusionThe use of xenogeneic collagen matrix in RT2 recessions was effective for recession reduction to those obtained using autologous grafts; with the advantage that the duration of surgery and patient morbidity decreased. Therefore, xenogeneic collagen matrix in RT2 recessions could be an alternative to autologous grafts.Clinical relevanceThe use of xenogeneic collagen matrix decreases the surgery time and patient morbidity but connective tissue graft results in significantly better mean root coverage and complete root coverage. Xenogeneic collagen matrix can be used in the treatment of RT2 gingival recessions.Study registrationNCT 03344315.

(248) Analysis of Biomarkers (MMP-9, TIMP-1, Ki-67 and Type IV Collagen) in the Diagnosis of Prostate Diseases

Abstract Introduction The combination of inflammatory, benign and malignant processes in the prostate reduces the sensitivity, specificity and diagnostic accuracy of modern examination methods. The development of immunohistochemical methods (IHC) promotes the search for molecular markers that are diagnostically significant in various processes in the prostate. This will improve the detection of malignant neoplasms and reduce the number of "unnecessary" prostate biopsies. Objective to study in biopsies of patients with various prostate diseases the relationship of type 9 metalloproteinase (MMP-9) and its inhibitor (TIMP-1), the role of the Ki-67 protein proliferation level and the diagnostic value of the type IV collagen distribution indicator. Methods The study included 263 patients. They were divided into 4 groups: chronic prostatitis - 21 patients (mean age 65.2 ± 0.7 years), benign prostatic hyperplasia (BPH) - 65 patients (69.3 ± 1.1 years), BPH + prostatic intraepithelial neoplasia (PIN) - 59 patients (68.9 ± 0.9 years), prostate adenocarcinoma (PC) - 118 patients (68.2 ± 0.9 years). IHC was used to analyze type 9 metalloproteinase (MMP-9) and its inhibitor (TIMP-1), the role of the Ki-67 protein proliferation level and the diagnostic value of the type IV collagen distribution indicator. Results BPH is characterized by high production of MMP-9 (intensity 3 points), lack of proliferative activity of the secretory cell layer (Ki-67 less than 3.24 ± 0.61%) and degradation of collagen IV of the basement membranes, which associated with a compensatory increase in the activity of the inhibitor of matrix metalloproteinases TIMP-1 (intensity 3 points). In prostate cancer, as neoplastic changes increase, the production of the Ki-67 protein significantly increases, reflecting the proliferative activity of the secretory cell layer (from 5.3 + 1.1% to 17.2 + 9.4%). With an increase in the Gleason number over the 3rd grade, collagen fibers around the tumor cells completely disappear. The production of MMP-9 type in adenocarcinomas is significantly reduced and the destruction of collagen IV of the basement membrane of glandular structures and stroma occurs due to a progressive (up to an intensity of 0.8 + 0.4 points as anaplastic changes increase) decrease in the production of the inhibitor of matrix metalloproteinases TIMP-1 (p≤0.05). High production of MMR-9 in BPH is blocked by a high content of the TIMP-1 inhibitor. In adenocarcinomas, the production of MMP-9 is significantly reduced, but this protease destroys collagen IV of the basement membrane of glandular structures due to the absence or weak production of TIMP-1, which blocks the proteolytic effect of this enzyme on connective tissue collagen. The low production of the TIMP-1 inhibitor in adenocarcinomas can also explain the invasive properties of these tumors. Conclusions Comprehensive analysis of biomarkers in IHC can improve the diagnosis of prostate cancer and reduce the number of "unnecessary" biopsies. Disclosure No.

The Effects of Massage with Topical Agents (Mepiform Ultra Scar Gel or Scarnos Gel) on Scar Tissue Thickness and Fibroblast Proliferation in Rats

Background: Keloid and hypertrophic scars are prominent scars that are excessively repaired with upregulated synthesis, deposition, and accumulation of collagen. Topical agents are used to reduce inflammation and fibrotic changes via reduced transepithelial water loss. Increased mechanical pressure applied through scar massage can accelerate scar maturation by inducing fibroblast proliferation, which enhances the remodeling of connective tissue matrices and collagen degradation.Methods: This study comparatively analyzed the effectiveness of topical agents applied to post-burn wounds on the dorsal surface of Sprague-Dawley rats through twice-daily massaging. Postoperative histological analysis of the tissues was performed after surgical en bloc removal of the treatment area at 4, 10, and 16 weeks.Results: Histological analyses revealed larger amounts of fibroblasts in Mepiform and Scarnos gel-treated tissue than in Vaseline-treated tissue. Granulation was prevented in scars treated with the topical agents.Conclusion: Mepiform Ultra scar gel and Scarnos gel, accompanied by massaging, may be effective anti-scarring topical agents to alleviate contact burn scars in Sprague-Dawley rats.

Highly specific imaging of pathological type I collagen in connective tissues by dual SERS peptide probes

The precise detection of pathological type I collagen is critical for diagnosing and prognosticating various diseases such as fibrosis and tumors. Surface-enhanced Raman scattering (SERS) is emerging as a promising high-resolution imaging technique, yet the intricate process of covalently modifying antibodies often results in SERS probes with diminished selectivity. We herein for the first time report the development of dual SERS peptide probes, Ag@R1@PCTP (S-P) and Ag@R2@ICTP (S-I), which integrate collagen targeting peptides (CTPs), Raman reporters, and silver nanoparticles (Ag NPs) for specific imaging of pathological type I collagen in connective tissues. The S-P/S-I dual SERS peptide probes displayed distinct peaks at 2154 cm−1 and 2227 cm−1 in the Raman silent region with no overlap, showcasing their capability for multiplex imaging free from cross-interference. The SERS peptide probe S-P is specifically tuned to target pathological collagen, excluding intact collagen, while S-I effectively recognizes type I collagen in both its pathological and healthy states. The S-P/S-I probes have exhibited outstanding effectiveness in identifying pathological type I collagen across various connective tissues and have been successfully applied for imaging this collagen in a variety of cancer tissues. The innovative dual SERS peptide probes offer a highly specific and robust tool for imaging pathological type I collagen in tissues, presenting significant potential for the accurate diagnosis and treatment of a range of collagen-related diseases.

(197) The Impact of Connective Tissue Disorders on Rates of Penile Fracture and Peyronie’s Disease

Abstract Introduction Ehlers Danlos Syndrome (EDS) and collagen-related hypermobility disorders are well known to cause frequent joint dislocations, easy bruising, and poor wound healing. Marfan syndrome is caused by a mutation in the protein fibrillin, a key component of collagen and elastin connective tissue, whose dysfunction causes the marfanoid phenotype. Currently, little is known regarding the impact of connective tissue disorders on men’s sexual health. As a first step in exploring this association, we investigated two physical phenomena that may be impacted by collagen disorders, penile fractures and Peyronie’s disease. Objective This study aims to investigate rates of penile fracture and Peyronie’s disease in EDS and Marfan syndrome patients. Methods A multi-center, international, electronic health record network (TriNetX) was queried to identify adult male patients with or without EDS between 1993 and 2023 using ICD-10 codes Q79.6, Q79.60, and Q79.62 specific for EDS and EDS related hypermobility syndrome. TriNetX was separately queried for Marfan syndrome using ICD-10 code Q87.4. Incidence and prevalence of penile fractures and Peyronie’s disease were compared between patients with and without the diseases of interest. Penile fractures were identified using ICD-10 codes S39.840, S39.84A, S39.84S, S39.84D. Peyronie’s disease was defined using ICD-10 code N48.6. Odds ratios were generated with 95% confidence intervals. All statistical analyses were conducted within the TriNetX platform. Results Data was queried from 82 healthcare organizations to yield 11,937 EDS patients, and from 101 healthcare organizations to yield 13,705 Marfan syndrome patients. 109 healthcare organizations contributed data to yield a control group of 63,790,628 men. Men with EDS had a 25 times higher rate of penile fractures (0.09%) compared to men without EDS (OR 25; 95% CI [13.8-47.8]). Men with Marfan’s syndrome had a 23 times higher rate of penile fractures (0.08%) compared to men without Marfan’s syndrome (OR 23; 95% CI [12.4-43.1]). Men with Marfan’s syndrome also had a significantly higher incidence of Peyronie’s disease compared to men without Marfan’s syndrome (OR 1.9; 95% CI [1.2-3]). The difference in rates of Peyronie’s disease was not significantly higher for men with EDS (OR 1.4; 95% CI [0.8 – 2.5]). Conclusions Patients with EDS and Marfan syndrome are at an over 20 times higher risk for penile fracture. These increased risks may be due to EDS’s and Marfan syndrome’s effects on collagen and fibrillin/elastic fibers, respectively, which are significant structural components of the corpora cavernosa. Patients with these syndromes should be aware of this increased risk and further research is needed to establish how to council these patients about taking appropriate precautions. Disclosure No.

Fluvoxamine supresses transforming growth factor beta‐2‐induced fibrotic changes in trabecular meshwork cells

Aims/Purpose: Primary open‐angle glaucoma is the leading cause of blindness worldwide, but no definitive treatment is available. Fibrotic‐like changes of trabecular meshwork (TM) results in increased stiffness and impaired outflow leading to increased IOP. Transforming growth factor‐β2 (TGF‐β2), a main driver of eye fibrosis, is increased in the aqueous humour of glaucoma patients. Recently, we proved that the specific sigma‐1 receptor agonist fluvoxamine (FLU) is antifibrotic in the kidney but its role in the TM is unknown. Therefore, we investigated the effects of FLU on the TGF‐β2‐induced fibrotic response in TM cells.Methods: Fibrosis of the AS was induced in C57BL/6J mice by intracameral TGF‐β2 injection. AS were collected and F‐actin was visualized with Phalloidin and detected with confocal microscopy. The effect of FLU on TGF‐β2‐induced cell proliferation, cytoskeletal rearrangement, and fibrosis‐related proteins was investigated by MTT (3‐[4,5‐dimethylthiazol‐2‐yl]‐2,5 diphenyl tetrazolium bromide) assay, Phalloidin staining and Western blotting, respectively.Results: TGF‐β2 injection resulted in F‐actin enhancement in vivo, indicating fibrotic‐like changes. In vitro, TGF‐β2 induced cytoskeletal rearrangement with increased F‐actin‐ bundles and clumps formation (p < 0.001) which were reduced by FLU (p < 0.001). Furthermore, FLU also inhibited cell proliferation (p < 0.05) in TGF‐β2‐treated cells. The profibrotic protein levels of connective tissue growth factor, fibronectin and collagen type IV were elevated by TGF‐β2 treatment (p < 0.001), while FLU prevented the elevation of these elements (p < 0.01; p < 0.001). FLU also hindered the TGF‐β2 caused decrease of Matrix Metalloproteinase‐2 (p < 0.05) thus facilitating ECM degradation.Conclusions: FLU may reduce the fibrotic response of trabecular meshwork and could be a potential candidate for the treatment of fibrosis‐induced ocular hypertension.Grants: OTKA‐ K135398, LP2021‐3/2021, TKP2021‐EGA‐24, Stipendium Hungaricum.

Immune Cell Profiling in the Ovarian Stroma of a Viviparous Fish during the Breeding Season: A Histological and Immunohistochemical Investigation

The molly fish is a member of viviparous teleosts that are characterized by the fusion of the right and left ovaries during their early embryonic development. This fusion results in a singular and saccular ovary, where the germinal epithelium lines the internal lumen. The present study aimed to identify the immune cells in the ovarian stroma of Molly fish during the breeding season using histological and immunohistochemical analysis. Histological examination of the ovaries displayed oocytes at all different stages of development and degeneration. The ovocoel, a lymph-filled space, remains in the center of the ovary and branches posteriorly, creating the lumen of the gonoduct. The ovarian wall is composed of three layers: the mesothelium, tunica albuginea, and germinal epithelium. The developing ova were held together by the stroma, which consisted of vascular collagenous connective tissue clustered with immune cells. Immunohistochemical analysis revealed the presence of clusters of macrophages expressing APG5, IL-1β, TGF-β, S100, NF-κB, CD68, Iba-1, and Ach. Monocytes demonstrated positive immunoreactivity for both APG5 and IL-1β, whereas dendritic cells expressed only APG5. Furthermore, rodlet cells exhibited immunoreactivity for S100 protein, IL-1β, NF-κB, CD68, Nrf2, Ach, myostatin, SOX9, and Iba-1. In contrast, stem cells displayed immunoreactivity for Nrf2, myostatin, and SOX9. In conclusion, the ovarian stroma of Molly fish demonstrated a notable presence of immune cells, indicating their active involvement in immune reactions.

In-silico and in-vitro screening of Asiatic acid and Asiaticoside A against Cathepsin S enzyme

BackgroundAtherosclerosis is a form of cardiovascular disease that affects the endothelium of the blood vessel. Series of events are involved in the pathophysiology of this disease which includes the breaking down of the connective tissue elastin and collagen responsible for the tensile strength of the arterial wall by proteolytic enzyme. One of these enzymes called Cathepsin S (CatS) is upregulated in the progression of the disease and its inhibition has been proposed to be a promising pharmacological target to improve the prognosis of the disease condition. Asiatic acid and asiaticoside A are both pentacyclic triterpenoids isolated from Centella asiatica. Their use in treating various cardiovascular diseases has been reported.MethodsIn this study through in silico and in vitro methods, the pharmacokinetic properties, residue interaction, and inhibitory activities of these compounds were checked against the CatS enzyme. The SwissADME online package and the ToxTree 3.01 version of the offline software were used to determine the physicochemical properties of the compounds.ResultAsiatic acid reported no violation of the Lipinski rule while asiaticoside A violated the rule with regards to its molecular structure and size. The molecular docking was done using Molecular Operating Environment (MOE) and the S-score of − 7.25988, − 7.08466, and − 4.147913 Kcal/mol were recorded for LY300328, asiaticoside A, and asiatic acid respectively. Asiaticoside A has a docking score value (− 7.08466Kcal/mol) close to the co-crystallize compound. Apart from the close docking score, the amino acid residue glycine69 and asparagine163 both interact with the co-crystallized compound and asiaticoside A. The in vitro result clearly shows the inhibitory effect of asiaticoside and asiatic acid. Asiaticoside A has an inhibitory value of about 40% and asiatic acid has an inhibitory value of about 20%.ConclusionThis clearly shows that asiaticoside will be a better drug candidate than asiatic acid in inhibiting the CatS enzyme for the purpose of improving the outcome of atherosclerosis. However, certain modifications need to be made to the structural make-up of asiaticoside A to improve its pharmacokinetics properties.

Xenogeneic collagen matrix vs. connective tissue graft for the treatment of multiple gingival recession: a systematic review and meta-analysis.

The aim of this systematic review (SR) compared the effect of xenogeneic collagen matrix (XCM) vs. connective tissue graft (CTG) for the treatment of multiple gingival recession (MGR) Miller Class I and II or Cairo type I. Five databases were searched up to August 2022 for randomized clinical trials (RCTs) comparing the clinical effects of XCM vs. CTG in the treatment of MGR. The random effects model of mean differences was used to determine reduction of gingival recession (GR), gain in keratinized tissue width (KTW), gain in gingival thickness (GT) and gain in clinical attachment level (CAL). The risk ratio was used to complete root coverage (CRC) at 6 and 12months. 10 RCTs, representing 1095 and 649 GR at 6 and 12months, respectively, were included in this SR. The meta-analysis showed no statistically significant difference in GR reduction, KTW gain GT gain or CAL gain between groups at 6months. However, at 12months of follow-up, differences favoring the control group were observed (p < 0.05). CRC was significantly higher in the CTG group at 6 and 12months. Regarding dentine hypersensitivity (DH), no statistically significant differences were found between groups at 6 and 12months of follow-up (p < 0.05). At 12months, CTG showed significantly superior clinical results in the treatment of MGR: however, this difference was not observed in the decrease of DH.

Effects of L-ascorbic acid (C6H8O6: Vit-C) on collagen amino acids: DFT study.

Vitamin C plays a very important role in the repair of connective tissue, especially for sports whose training causes the most damage to this tissue. Therefore, many people believe that L-ascorbic acid (C6H8O6: vitamin C) reduces the recovery time between sports exercises. The most abundant form of structural protein in the body is collagen. Collagen is characterized by a high concentration of the three amino acids glycine (Gly), proline (Pro), and hydroxyproline (Hyp), which creates its characteristic triple helix structure. Therefore, in this study, the effect of vitamin C presence on the sequence, interaction, and orientation of amino acids for collagen formation is investigated using computational simulation. This study aimed to investigate the mechanism of action of vitamin C in terms of thermodynamics and structure of the reaction. The calculations are performed using density function theory (DFT) by the base set of B3LYP/6-311++G (p,d). The results show that the presence of vitamin C is effective in the formation of collagen protein for this interaction and the mechanism of amino acid sequence (Gly-Hyp-Pro) is better in the formation of collagen protein in the presence of vitamin C. The presence of Vit-C in the formation and direction of hydroxyproline (Hyp) causes its separation from the prolyl 5-hydroxylase enzyme. In the absence of vitamin C, the reaction stops at this stage and proline cannot be converted into hydroxyproline. The computational data shows vitamin C prevents unwanted interactions and directs amino acid reactions to repair connective tissue (collagen). Therefore, vitamin C acts as a cofactor in the Prolyl 5-Hydroxylase enzyme and causes it to convert proline to hydroxyl.

Orofacial Manifestations in a Middle-Aged Woman with Cowden Syndrome: A Case Image.

A 56-year-old Brazilian woman sought dental care, presenting with multiple asymptomatic papillomatous lesions with a coalescent pattern and intermingled cobblestone-like clefts along the alveolar ridge and marginal and attached gingivae. Multiple whitish papules were also observed on the face, neck, and limbs. Incisional biopsies of these lesions were performed. Microscopically, the skin lesion revealed epithelial clear cells and intraepithelial keratinization with areas of orthokeratosis, while the gingival lesions showed a parakeratinized stratified squamous epithelium with collagenous connective tissue. These features were consistent with those of a trichilemmoma and fibroepithelial hyperplasia, respectively. This article illustrates a case of Cowden syndrome (CS), a rare multisystem genetic condition in which both cutaneous and mucosal tissues were affected. Fewer than 40 cases of CS with oral involvement affecting middle-aged adults have been documented hitherto.