Tissue Growth Factor Research Articles

Novel regulation of kidney function by gastrointestinal hormones

The control of whole-body energy metabolism involves intricate crosstalk between multiple organ systems including the gut and the brain that play well-recognized roles in this process. However, the important contributions and mechanisms by other organs are poorly understood. Errors in metabolic regulation can lead to human metabolic diseases including diabetes and obesity that affect 75% of the population and can lead to significant comorbidities. Improving our mechanistic understanding of multi-organ metabolic crosstalk in health and disease is essential for developing highly effcient and new therapeutic approaches. Macula densa (MD) cells in the kidney are chief regulators of renal blood flow and tissue remodeling. We hypothesized that the kidney is a major player in body metabolism via a new mechanism of gut-kidney crosstalk that involves the gut hormones gastrin and cholecystokinine (CCK) activating intrarenal, MD-specific mechanisms to increase blood flow and filtration, progenitor cells and tissue growth factors. High-resolution MD bulk and single-cell transcriptome analysis was performed from freshly isolated, enriched MD cells using MD-GFP mouse kidneys. Single cell MD Ca2+ signaling was analyzed in vivo using intravital multiphoton microscopy (MPM) of Sox2-GT mice that express the ratiometric Ca2+ reporter GCaMP6f/tdTomato in all renal cells. The altered expression of MD specific markers in response to gastrin/CCK8 stimuli was studied using western blot and the novel MDgeo immortalized cell line cultured in vitro. RNA sequencing and transcriptome analysis identified the gastrin/CCK receptor Cckbr as one of the highest expressed genes in MD versus control kidney cells. Intravital multiphoton imaging of Sox2-GCaMP6 reporter mouse kidneys revealed robust and entirely MD cell-specific calcium signaling in response to systemic gastrin/CCK injection. Using the novel MDgeo immortalized cell line cultured in vitro, CCK8 treatment led to 3-fold increases in MD cell synthesis of Prostaglandin E2 (PGE2)-generating COX2 and CCN1 that are established mediators of MD-induced glomerular hyperfiltration and tissue remodeling, respectively. Chronic treatment of Ren1d-Confetti mice with Darunavir, an HIV protease inhibitor significantly increased plasma CCK levels, Ren+ mesenchymal progenitor cell number and clonality, renal cell proliferation and tissue hypertrophy. This study uncovered new regulatory mechanisms of systemic metabolism especially the role of MD cells in the kidney. Repurposing the commonly used HIV drug Darunavir for kidney and metabolic diseases may provide therapeutic benefit for millions of patients worldwide. DK064234, DK123564, DK135290. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Toward the Development of a Tissue Engineered Gradient Utilizing CRISPR-Guided Gene Modulation.

Cellular, compositional, and mechanical gradients are found throughout biological tissues, especially in transition zones between tissue types. Yet, strategies to engineer such gradients have proven difficult due to the complex nature of these tissues. Current strategies for tissue engineering complex gradients often utilize stem cells; however, these multipotent cells require direction from environmental cues, which can be difficult to control both in vitro and in vivo. In this study, we utilize clustered regularly-interspaced short palindromic repeats (CRISPR)-guided gene modulation to direct the differentiation of multipotent adipose-derived stem cells (ASCs) to demonstrate the effectiveness of CRISPR-engineered cells in tissue engineering applications. Specifically, we screen CRISPR-interference (CRISPRi) constructs targeting the promotors of selected osteogenic inhibitors and demonstrate that ASC osteogenic differentiation and mineral deposition can be regulated with CRISPRi targeting of Noggin without the use of exogenous growth factors in tissue engineered constructs. As a proof of concept, we combine three technologies developed out of our laboratories to demonstrate the controlled deposition of these engineered cells in a gradient with CRISPR-activation multiplex-engineered aggrecan/collagen type-II-chondrogenic ASCs on a high density anisotropic type I collagen construct to create a cell and tissue gradient similar to the fibrocartilage-to-mineralized-fibrocartilage gradient in the enthesis. Our results display the promise of CRISPR-engineered ASCs to produce tissue gradients, similar to what is observed in native tissue.

(023) Mechanical Therapy After Inflatable Penile Prosthesis: A Scoping Review

Abstract Introduction While inflatable penile prosthesis (IPP) allows reliable axial rigidity, common complaints perceived are shorter penis and reduced glans engorgement. Pre-operative factors, such as vasculogenic erectile dysfunction (ED) or corporoglandular morphology, provide a convenient explanation. However, post-operative factors are more recently being interrogated. Post-IPP mechanical therapy protocols, like vacuum erection devices (VED), are being instituted to ameliorate these complaints. The physiological rationale is to improve perfusion of residual erectile tissues and limit cavernous fibrosis/pseudo-capsule formation. This has been hypothetically explained by upregulating endothelial nitrous oxide synthase release, decreasing hypoxia inducible factor and tissue growth factor 1. Post-IPP maximal inflation protocols similarly resulted in a more capacious and compliant tunica albuginea and capsule. Objective To assess the utility of post-IPP mechanical therapy in addressing common post-operative complaints and enhance prosthetic erection. Methods Intervention: Post-op mechanical therapy Population: Men with IPP for ED, with/without Peyronie’s disease (PD) Inclusion Criteria: Articles published in English with full-text available Search Strategy: Literature search was performed in June 2023 using Medline, PubMed, Embase and Scopus. PubMed search included “Mesh” protocols and “free text” to correctly capture study population. Articles relating to the intervention were identified with free text search searches (“VED,” “vacuum erection device,” “vacuum constriction device,” “penis pump,” “mechanical therapy,” or “rehabilitation”). Review Type & Analysis: While a systematic model was initially adopted, a scoping model was ultimately chosen given the limited available literature & variable outcome measures. In this way, we sought to identify knowledge gaps and scope of current literature to guide further studies in this field. Higher-powered analyses were not possible due to inter-study outcome measure variability. Results Initial search strategy retrieved 81 non-duplicate studies with 30 selected for initial screening after search limitations. Ultimately, 5 satisfied inclusion criteria after full-text review and were divided into 3 categories according to post-IPP protocol employed: Early activation (1), Maximal inflation (2,3) and VED (4,5) (Table 1). A common theme encountered was that mechanical therapy improved post-IPP penile length and girth, compared to immediate post-operative measurements. Only one study used a control group who had their device activated for the first time after 4 weeks. The intervention employed in this study was device activation immediately after surgery, left 80% inflated and deflated at 3 weeks, then full erection at night for 3 further weeks. Compared to the control, at maximum inflation, penile length mean was 3.28cm longer and circumference was 1cm greater. In each of these studies, the differences reported were statistically significant. This was also associated with an increased number of pumps required to achieve full inflation over time, irrespective of mechanical protocol used. Articles that reported IIEF-5 and EDITS scores demonstrated a statistically significant improvement over time with post-op intervention. Conclusions Within limits of the included studies, post-implantation mechanical therapy appears to be useful in improving penile dimensions and patient satisfaction over time. The correlation with increasing number of pumps implies that compliance of the tunica albuginea and residual cavernous tissue may play a role in the pathogenesis of shorter penis post-IPP. Further high-powered studies in this area are warranted. Disclosure No.

(312) Manhandling: Post-IPP Mechanical Therapy – A Scoping Review

Abstract Introduction While inflatable penile prosthesis (IPP) allows reliable axial rigidity, common complaints are perceived shorter penis and reduced glans engorgement. Pre-operative factors, such as vasculogenic erectile dysfunction (ED) or corporoglandular morphology, provide a convenient explanation. However, post-operative factors are more recently being interrogated. Post-IPP mechanical therapy protocols, like vacuum erection devices (VED), are being instituted to ameliorate these complaints. The physiological rationale is to improve perfusion of the residual erectile tissues and limit cavernous fibrosis or pseudo-capsule formation. This has been hypothetically explained by upregulating release of endothelial nitrous oxide synthase, decreasing hypoxia inducible factor  and tissue growth factor □1. Post-IPP maximal inflation protocols may similarly result in a more capacious and compliant tunica albuginea and capsule. Objective To assess the utility of post-IPP mechanical therapy in addressing common post-operative complaints and enhance prosthetic erection. Methods Intervention: Post-op mechanical therapy Population: Men with IPP for ED, with/without Peyronie’s disease (PD) Inclusion Criteria: Articles published in English with full-text available Search Strategy: Literature search was performed in June 2023 using Medline, PubMed, Embase and Scopus. PubMed search included both “Mesh” protocols and “free text” to capture the correct study population. Articles relating to the intervention were identified with free text search searches (“VED,” “vacuum erection device,” “vacuum constriction device,” “vacuum,” “penis pump,” “mechanical therapy,” or “rehabilitation”). Review Type & Analysis: While a systematic model was initially adopted, a scoping model was ultimately chosen given the limited available literature and variable outcome measures. In this way, we sought to identify knowledge gaps and the scope of current literature to guide further studies in this field. Higher-powered analyses were not possible due to inter-study outcome measure variability. Results Initial search strategy retrieved 81 non-duplicate studies with 30 selected for initial screening after search limitations. Ultimately, 5 satisfied inclusion criteria after full-text review and were divided into 3 categories relating to the post-IPP protocol employed: Early activation (1), Maximal inflation (2,3) and VED (4,5) (Table 1). A common theme encountered was that mechanical therapy improved post-IPP penile length and girth, compared to immediate post-operative measurements. Only one study used a control group who had their device activated for the first time after 4 weeks. The intervention employed in this study was device activation immediately after surgery, left 80% inflated and deflated at 2-3 weeks, then full erection at night for 3-4 further weeks. Compared to the control, at maximum inflation, penile length mean was 3.28 cm longer and circumference was 1 cm greater. In each of these studies, the differences reported were statistically significant. This was also associated with an increased number of pumps required to achieve full inflation over time, irrespective of mechanical protocol used. Articles that reported IIEF-5 and EDITS scores also demonstrated a statistically significant improvement over time with post-op intervention. Conclusions Within limits of the included studies, post-implantation mechanical therapy appears to be useful in improving penile dimensions and patient satisfaction over time. The correlation with increasing number of pumps implies that compliance of the tunica albuginea and residual cavernous tissue may play a role in the pathogenesis of shorter penis post-IPP. Further high-powered studies in this area are warranted. Disclosure No.

The inhibition of YAP Signaling Prevents Chronic Biliary Fibrosis in the Abcb4-/- Model by Modulation of Hepatic Stellate Cell and Bile Duct Epithelium Cell Pathophysiology.

Primary sclerosing cholangitis (PSC) represents a chronic liver disease characterized by poor prognosis and lacking causal treatment options. Yes-associated protein (YAP) functions as a critical mediator of fibrogenesis; however, its therapeutic potential in chronic biliary diseases such as PSC remains unestablished. The objective of this study is to elucidate the possible significance of YAP inhibition in biliary fibrosis by examining the pathophysiology of hepatic stellate cells (HSC) and biliary epithelial cells (BEC). Human liver tissue samples from PSC patients were analyzed to assess the expression of YAP/connective tissue growth factor (CTGF) relative to non-fibrotic control samples. The pathophysiological relevance of YAP/CTGF in HSC and BEC was investigated in primary human HSC (phHSC), LX-2, H69, and TFK-1 cell lines through siRNA or pharmacological inhibition utilizing verteporfin (VP) and metformin (MF). The Abcb4-/- mouse model was employed to evaluate the protective effects of pharmacological YAP inhibition. Hanging droplet and 3D matrigel culture techniques were utilized to investigate YAP expression and activation status of phHSC under various physical conditions. YAP/CTGF upregulation was observed in PSC patients. Silencing YAP/CTGF led to inhibition of phHSC activation and reduced contractility of LX-2 cells, as well as suppression of epithelial-mesenchymal transition (EMT) in H69 cells and proliferation of TFK-1 cells. Pharmacological inhibition of YAP mitigated chronic liver fibrosis in vivo and diminished ductular reaction and EMT. YAP expression in phHSC was effectively modulated by altering extracellular stiffness, highlighting YAP's role as a mechanotransducer. In conclusion, YAP regulates the activation of HSC and EMT in BEC, thereby functioning as a checkpoint of fibrogenesis in chronic cholestasis. Both VP and MF demonstrate effectiveness as YAP inhibitors, capable of inhibiting biliary fibrosis. These findings suggest that VP and MF warrant further investigation as potential therapeutic options for the treatment of PSC.

Sustainable Treatment of Oral Traumatic Ulcers with Licorice Containing Hydrogels: Integrating Computational Modeling, Quality by Design, Green Synthesis, and Molecular Biological Evaluation.

The urge to implement innovative approaches that align with eco-friendly practices and hold promise for enhancing oral health while promoting environmental sustainability has been increasing. This current work aims to develop a sustainable treatment for oral traumatic ulcers using licorice-based hydrogels (LHGs) containing hydroxyethyl cellulose (HEC) as the green gelling agent. Licorice root aqueous extract was phytochemically profiled using UPLC-ESI-MS/MS. Forty-three compounds were detected, with Glycyrrhizic acid being the major component of the extract (34.85 ± 2.77%). By implementing a Quality by Design (QbD) approach, the study investigates the effects of different licorice extract and HEC concentrations on key variables such as pH and viscosity of the prepared formulations, ulcer and wound healing scores, and tissue growth factors via a Full Factorial Experimental Design. The LHGs exhibited desirable consistency, spreadability, and clarity. Statistical analysis, employing an ANOVA test, revealed the high significance of the constructed models with the licorice concentration being the key independent factor affecting all dependent outputs. The pH as well as the viscosity of the prepared LHGs were positively influenced by licorice extract concentration, with higher concentrations leading to increased alkalinity and viscosity. Rheological behavior analysis revealed a pseudoplastic flow with demonstrated thixotropy which is advantageous for application and prolongation of residence time. The wound healing process was assessed through ulcer size, traumatic ulcer healing score (UHS), collagen-1 expression (COL-1), growth factors (EGF, VEGF), pro-inflammatory markers (TNF-α), wound healing score (WHS). LHGs prepared using higher levels of both factors, 30% dried licorice root extract and 4% HEC, demonstrated enhanced wound healing, elevated growth factor expression of 66.67% and 23.24%, respectively, and 88% reduced inflammation compared to the control group, indicating their potential in expediting oral ulcer recovery. Overall, these findings highlight the promising role of green licorice-based hydrogels in promoting sustainable oral mucosal healing.

Ezrin promotes esophageal squamous cell carcinoma progression via the Hippo signaling pathway.

The aim of this study was to analyze the role of Ezrin in esophageal squamous cell carcinoma (ESCC) and investigate potential therapeutic targets for ESCC by interfering with Ezrin expression. Bioinformatics analysis revealed that Ezrin expression differed significantly among patients with different clinical stage ESCC. Moreover, there was a significant correlation between Ezrin and yes-associated protein/connective tissue growth factor (YAP1/CTGF) levels in esophageal cancer. Sixty paraffin-embedded ESCC tissue samples were examined and Ezrin and YAP1/CTGF levels were determined using immunohistochemistry. The positive expression rates of Ezrin and YAP1/CTGF were significantly lower in adjacent tissues than in ESCC tissues. Furthermore, knockdown of Ezrin expression inhibited colony formation and reduced cell migration and invasion. Compared with control ESCC cells, protein expression levels of YAP1 and CTGF were significantly downregulated in cells with Ezrin knocked down. We conclude that Ezrin may be involved in ESCC progression through the Hippo signaling pathway.

Airway epithelial cells mount an early response to mycobacterial infection.

Lung epithelial cells represent the first line of host defence against foreign inhaled components, including respiratory pathogens. Their responses to these exposures may direct subsequent immune activation to these pathogens. The epithelial response to mycobacterial infections is not well characterized and may provide clues to why some mycobacterial infections are cleared, while others are persistent and pathogenic. We have utilized an air-liquid interface model of human primary bronchial epithelial cells (ALI-PBEC) to investigate the epithelial response to infection with a variety of mycobacteria: Mycobacterium tuberculosis (Mtb), M. bovis (BCG), M. avium, and M. smegmatis. Airway epithelial cells were found to be infected by all four species, albeit at low frequencies. The proportion of infected epithelial cells was lowest for Mtb and highest for M. avium. Differential gene expression analysis revealed a common epithelial host response to mycobacteria, including upregulation of BIRC3, S100A8 and DEFB4, and downregulation of BPIFB1 at 48h post infection. Apical secretions contained predominantly pro-inflammatory cytokines, while basal secretions contained tissue growth factors and chemokines. Finally, we show that neutrophils were attracted to both apical and basal secretions of infected ALI-PBEC. Neutrophils were attracted in high numbers to apical secretions from PBEC infected with all mycobacteria, with the exception of secretions from M. avium-infected ALI-PBEC. Taken together, our results show that airway epithelial cells are differentially infected by mycobacteria, and react rapidly by upregulation of antimicrobials, and increased secretion of inflammatory cytokines and chemokines which directly attract neutrophils. Thus, the airway epithelium may be an important immunological component in controlling and regulating mycobacterial infections.

Module 4-Deficient CCN2/Connective Tissue Growth Factor Attenuates the Progression of Renal Fibrosis via Suppression of Focal Adhesion Kinase Phosphorylation in Tubular Epithelial Cells.

CCN2/connective tissue growth factor (CTGF) potentially serves as a therapeutic target for chronic kidney disease. Here we investigated CCN2 module-4, encoded by Ccn2 exon 5, through the generation of Ccn2 exon 5 knockout mice (Ex5-/- mice). To investigate renal fibrosis pathogenesis, Ex5-/- mice were employed to model unilateral ureteral obstruction (UUO), unilateral ischemic-reperfusion injury (UIRI), and 5/6 nephrectomy. Interstitial fibrosis was significantly attenuated in the Ex5-/- mice in the three models. Furthermore, phosphorylated focal adhesion kinase (FAK) levels in tubular epithelial cells were significantly lower in the kidneys of the UUO- and UIRI-Ex5-/- mice than those of the Ex5+/+ mice. Moreover, CCN2 module 4-mediated renal tubule FAK and promoted fibrosis. These findings indicate that CCN2 module-4-FAK pathway components will serve as therapeutic targets for effectively attenuating renal fibrosis.

Blocking CCN2 Reduces Established Palmar Neuromuscular Fibrosis and Improves Function Following Repetitive Overuse Injury.

The matricellular protein cell communication factor 2/connective tissue growth factor (CCN2/CTGF) is critical to development of neuromuscular fibrosis. Here, we tested whether anti-CCN2 antibody treatment will reduce established forepaw fibro-degenerative changes and improve function in a rat model of overuse injury. Adult female rats performed a high repetition high force (HRHF) task for 18 weeks. Tissues were collected from one subset after 18 wks (HRHF-Untreated). Two subsets were provided 6 wks of rest with concurrent treatment with anti-CCN2 (HRHF-Rest/anti-CCN2) or IgG (HRHF-Rest/IgG). Results were compared to IgG-treated Controls. Forepaw muscle fibrosis, neural fibrosis and entheseal damage were increased in HRHF-Untreated rats, compared to Controls, and changes were ameliorated in HRHF-Rest/anti-CCN2 rats. Anti-CCN2 treatment also reduced phosphorylated-β-catenin (pro-fibrotic protein) in muscles and distal bone/entheses complex, and increased CCN3 (anti-fibrotic) in the same tissues, compared to HRHF-Untreated rats. Grip strength declines and mechanical sensitivity observed in HRHF-Untreated improved with rest; grip strength improved further in HRHF-Rest/anti-CCN2. Grip strength declines correlated with muscle fibrosis, entheseal damage, extraneural fibrosis, and decreased nerve conduction velocity, while enhanced mechanical sensitivity (a pain-related behavior) correlated with extraneural fibrosis. These studies demonstrate that blocking CCN2 signaling reduces established forepaw neuromuscular fibrosis and entheseal damage, which improves forepaw function, following overuse injury.

The Wound Healing Potential of 2,3 Dimethylquinoxaline Hydrogel in Rat Excisional Wound Model

Background: 2,3-dimethylquinoxaline (DMQ) is a naturally occurring compound with documented antifungal activity. It showed also good in vitro physicochemical and pharmacokinetic characteristics.
 Methods: The wound healing activity of 1% DMQ hydrogel in healthy adult male Wister rats were evaluated using the excision wound model. On day 7, the mean percentage closure of the wound area was determined. The animals were sacrificed on day 7 and skin was isolated for histology research and assessment of some inflammatory & oxidative markers, hydroxyproline and tissue growth factor.
 Results and Discussion: Almost complete wound healing was observed after treatment with DMQ 1 % Jell for 7 days. The histological study confirmed marked attenuation of wound-induced histological changes. There was a marked reduction in TNF-α, IL-6 IL-β1 and NF-κB. levels. These data suggest the potential anti-inflammatory effect of DMQ.
 Conclusion: DMQ has potential skin wound healing ability likely due to its anti-inflammatory mechanism. Further study is needed to confirm these preliminary findings and explore the molecular mechanism

Experimental And Clinical Evidence of the Sulfide Balneotherapy Influence Efficacy on the Trophological and Regenerative Status: a Fundamental, Randomized Controlled Trial

INTRODUCTION. The therapeutic strategies for managing patients with comorbid conditions, such as arterial hypertension (AH) and prediabetes, are widely adopting non-drug methods along with the recommended medications. AIM. To evaluate the effect on fat distribution and humoral status of sulfide balneotherapy in the conditions of the experiment on white rats and clinical study of AH and prediabetes patients. MATERIAL AND METHODS. The study was of a clinical and experimental nature and consisted of 2 phases – experimental and clinical. The experimental study was carried out on obese, non-pedigreed white rats. Animals were exposed to sulfide baths and a combination of sulfide applications and baths. In the clinical phase of the study, patients with AH and prediabetes were included. All patients included in the present study received standardised drug therapy according to clinical guidelines, and patients in the observation group additionally received combined sulphide balneotherapy (Suksun lake sulphide applications and baths) for 14 days. The study program included assessment of weight, waist circumference (WC), body mass index (BMI), assessment of serum concentrations of vascular endothelial growth factor (VEGF); tissue growth factor (TGF b), insulin and leptin, as well as determination of epicardial fat thickness (EFT). RESULTS AND DISCUSSION. The experimental conditions revealed lipolysis in adipocytes (omentum), as well as an increase in the number of hemocapillaries in all areas of the heart and the growth of CD34 expression. The results of the clinical stage of the study suggest redistribution of fat depots against the background of sulfide balneotherapy. There were no statistically significant differences in weight and BMI in the dynamics, but there was a significant decrease in OT and TEF in the group of patients who received balneotherapy, who also had a significant decrease in HEFR concentration, leptin and an increase in TEFR beta 1. CONCLUSION. The presence of lipolytic, regenerative and adipokine-regulating effects of sulfide balneotherapy was demonstrated in the experiment and clinical study.

Angiogenesis Assays for the Analysis of CCN Proteins.

Angiogenesis, the process of generating new blood vessels from an existing vasculature, is essential in normal developmental processes such as endochondral ossification and in numerous kinds of pathogenesis including tumor growth. A part from the actin of angiogenic factor or antiangiogenic factor, it is still unknown at which stage of the angiogenic cascade these agents affect angiogenesis. Here, we describe methods for the use of cellular communication network factor/connective tissue growth factor (CTGF/CCN2) and CCN2-neutralizing antibody in the currently used principal angiogenesis assays, including those in vitro ones for the proliferation, migration, adhesion, and tube formation of endothelial cells and in vivo assays such as those utilizing type I collagen implantation and the chick chorioallantoic membrane (CAM). In addition, we introduce an autofluorescence imaging of blood vessels in the subcutaneous tumor xenograft mouse model. These assays can be applied to studies on roles of CCN proteins in tumor metastasis and development of treatment strategies targeting CCN proteins.

Mouse Models of Tumor Bone Metastasis and Invasion for Studying CCN Proteins.

Bone metastasis and bone destruction are common occurrences in human malignancies, including breast, prostate, and lung cancer, and are associated with a high morbidity rate because of intractable bone pain, pathological fractures, hypercalcemia, and nerve compression. Animal models of bone metastasis and bone destruction are important tools to investigate the pathogenesis and develop treatment strategies. However, there are few models of spontaneous bone metastasis despite the fact that animals often spontaneously develop cancer. Here, we describe methods for developing a mouse model of breast cancer bone metastasis achieved by injection of MDA-MB-231 breast cancer cells into the left cardiac ventricle. In addition, we introduce mouse model of the bone destruction by injection of SAS oral squamous cell carcinoma cells into the bone marrow space of the right tibial metaphysis. These assays can be applied to studies on roles of cellular communication network factor/connective tissue growth factor (CTGF/CCN2) protein in tumor metastasis and development of treatment strategies targeting CCN proteins.

Complement 3a Mediates CCN2/CTGF in Human Retinal Pigment Epithelial Cells.

Background Complement 3 (C3) is the crucial component of the complement cascade when retina was exposed to external stimulus. Cellular communication network 2/connective tissue growth factor (CCN2/CTGF) is important in response of retinal stress and a fulcrum for angiogenesis and fibrosis scar formation. Our study aims to explore the interaction between C3 and CCN2/CTGF via bioinformatics analyses and in vitro cell experiments. Methods The GSE dataset was selected to analyse the chemokine expression in human retinal pigment epithelium (ARPE-19) cells under stimulus. Then, RPE cells were further transfected with or without C3 siRNA, followed by C3a (0.1 μM or 0.3 μM) for 24, 48, and 72 hours. Reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) were used to measure CCN2/CTGF mRNA and protein levels. Results The GSE36331 revealed C3 expression was significantly elevated in RPE under stimulus. Compared with negative control, CCN2/CTGF mRNA was increased with all types of C3a treatments, whereas a significant increase of protein level was only observed with high concentration of 0.3 μM C3a for a prolonged 72-hour time. Compared with nontransfected cells, significant reductions of CCN2/CTGF mRNA were observed in the C3 siRNA transfected cells with 0.3 μM C3a for 24, 48, and 72 hours, and a significant reduction of CCN2/CTGF protein was observed with 0.3 μM C3a for 48 hours. Conclusions C3 was elevated in RPE under environmental stimulus and long-term exposure to specified concentration of C3a increased CCN2/CTGF expression in RPE, which could be partially reversed by C3 siRNA.

Topical Biological Agents as Adjuncts to Improve Wound Healing in Chronic Diabetic Wounds: A Systematic Review of Clinical Evidence and Future Directions.

Diabetes is a leading chronic illness in the modern world and 19-34% develop chronic diabetic foot ulcers (DFUs) in their lifetime, often necessitating amputation. The reduction in tissue growth factors and resulting imbalance between proteolytic enzymes and their inhibitors, along with systemic factors impairing healing appear particularly important in chronic wounds. Growth factors applied topically have thus been suggested to be a non-invasive, safe, and cost-effective adjunct to improve wound healing and prevent complications.Comprehensive database searches of MEDLINE via PubMed, EMBASE, Cochrane, and ClinicalTrials.gov were performed to identify clinical evidence and ongoing trials. The risk of bias analysis included randomized controlled trials (RCTs) was performed using the Cochrane Risk of Bias 2.0 tool. We included randomized controlled trials that compared the use of a topical biologic growth factor-containing regimen to any other regimen. Primary outcomes of interest were time to wound closure, healing rate, and time. Secondary outcomes included the incidence of adverse events such as infection.A total of 41 trials from 1992-2020 were included in this review, with a total recorded 3,112 patients. Platelet-derived growth factors (PDGF) in the form of becaplermin gel are likely to reduce the time of closure, increase the incidence of wound closure, and complete wound healing. Human umbilical cord-related treatments, dehydrated human amnion and chorion allograft (dHACA), and hypothermically stored amniotic membrane (HSAM), consistently increased the rates and incidence of complete ulcer healing while reducing ulcer size and time to complete ulcer healing. Fibroblast growth factor-1 (FGF1) showed only a slight benefit in multiple studies regarding increasing complete ulcer healing rates and incidence while reducing ulcer size and time to complete ulcer healing, with a few studies showing no statistical difference from placebo. Platelet-rich fibrin (PRF) is consistent in reducing the time to complete ulcer healing and increasing wound healing rate but may not reduce ulcer size or increase the incidence of complete ulcer healing.Targeting the wound healing pathway via the extrinsic administration of growth factors is a promising option to augment wound healing in diabetic patients. Growth factors have also shown promise in specific subgroups of patients who are at risk of significantly impaired wound healing such as those with a history of secondary infection and vasculopathy. As diabetes impairs multiple stages of wound healing, combining growth factors in diabetic wound care may prove to be an area of interest. Evidence from this systematic literature review suggests that topical adjuncts probably reduce time to wound closure, reduce healing time, and increase the healing rate in patients with chronic DFUs.

Random Walk Algorithm-Based Computer Tomography (CT) Image Segmentation Analysis Effect of Spiriva Combined with Symbicort on Immunologic Function of Non-Small-Cell Lung Cancer.

The objective of this research was to explore the effect of the treatment regimen of Spiriva combined with Symbicort on the immune function of non-small-cell lung cancer (NSCLC) based on computed tomography (CT) imaging features. An automatic CT image segmentation algorithm (RW-CT) was constructed based on random walk (RW) and image segmentation technology. The image segmentation algorithm based on the Toboggan method (C-CT) was introduced to compare with the traditional RW algorithm. 60 subjects were divided into four groups: a Chinese combined with Western medicine group (treated with Spiriva combined with Symbicort, group C+W), a Chinese medicine group (treated with Spiriva, group C), a Western medicine group (treated with Symbicort, group W), and a model group for control (group M). The results show that the Dice coefficient of the RW-CT algorithm was obviously larger than that of the C-CT algorithm and the RW algorithm, while the Hausdorff distance (HD) of the RW-CT algorithm was much smaller than that of the other two algorithms (P < 0.05). The proportion of positive cells of hypoxia-inducible factor-1α (HIF-1α) in group C+W was the least (15%-23%), followed by the group W (21%-29%) and the group C (28%-37%), and that in the group M was the highest (39%-49%). There was a remarkable difference in the immunohistochemical scores (HIS) of vascular endothelial growth factor (VEGF) in the tumor tissues between group C+W and the group M (H = 14.200, P = 0.001), but there was no great difference from the group C and the group W (P > 0.05). There was a notable difference in the IHS of vascular endothelial factor recepto-2 (VEGFR-2) between the group C+W medication group and the group M (H = 12.800, P = 0.002), and there was no statistical difference between the group C and W (P > 0.05). In short, the RW-CT constructed based on RW was better than the traditional algorithms for CT image segmentation. The Spiriva combined with Symbicort could effectively inhibit the expression of VEGF, VEGFR-2, and HIF-1α in NSCLC and promote the immunologic function of the body.

Gender features of the chronic pain effect on some vascular endothelial growth factors and their skin receptors on the B16/F10 melanoma model.

e24061 Background: Patients with oncological pathology, especially with the progression of the disease, often have chronic pain[1]. Pain effects singly can cause dysfunction of the vascular system, which could be an additional risk of the tumor process progression. The aim of the study was to identify gender-specific chronic pain effects on some vascular endothelial growth factors and their skin receptors in the B16/F10 melanoma model. Methods: The study was conducted on mice of the C57BL/6 line (females, males; age 8 weeks, weight 21-22 g, n = 48, 12 animals in each group). Two weeks after the ligation of the sciatic nerve melanoma B16/F10 was transplanted under the skin in the back of the main groups of animals (I- females, II-males); control groups were held the same procedure without ligation (III-females, IV-males). The levels of VEGFA, sVEGFR1, VEGFC, sVEGFR3 were determined in intact skin by ELISA. All manipulations were carried out in accordance with the European Convention for the Protection of Vertebrates Used for Experimental and Other Scientific Purposes (ETS 123). Results: Chronic pain contributed to the accumulation of VEGF-A, sVEGF-R1, VEGF-C in the skin and the reduction of sVEGF-R3 in all animals: in females - 2.7 times, 2.9 times, 6.1 times and 6.1 times, accordingly, in males - 3.1 times, 3.7 times, 3.5 times and 7.2 times, accordingly. Consequently, in males the amplitude of fluctuations of VEGF and their receptors in the skin against the background of pain was higher than in females, except for VEGF-C, the amount of which increased in females more than in males, despite its identical initial level in the skin of animals of both sexes. As a result, the skin of males with pain contained more VEGF-A, sVEGF-R1 and sVEGF-R3, accordingly, by 1.4 times (p &lt; 0.05), 2.0 times and 1.9 times (p &lt; 0.05) and less VEGF-C - by 1.8 times (p &lt; 0.05). Conclusions: Chronic neuropathic pain had different effects on changes in the level of vascular endothelial growth factors and their receptors in intact skin in females and males of melanoma B16/F10 tumor carriers. The gender characteristics of the effect of chronic pain on neoangiogenesis may be associated with different rates of growth and development of the malignant process in male and female mice in the experiment.

Impact of Intragastric Balloon Placement on the Stomach Wall: A Prospective Cohort Study.

Endoscopic intragastric balloon (IGB) placement is a minimally invasive treatment for morbid obesity that is sometimes used as a preparatory step before surgical intervention. This study was performed to analyze the changes in the stomach wall induced by IGB placement, with particular emphasis on pathomorphology, inflammatory markers, and tissue growth factors. In total, 30 patients with morbid obesity were prospectively analyzed. A total of 16 patients with body mass index (BMI) ≥ 53kg/m2 underwent two-stage treatment comprising IGB placement followed by laparoscopic sleeve gastrectomy (LSG) (IGB group), while 14 patients underwent one-stage LSG (non-IGB group). The gastric specimens removed during LSG were examined. The two groups were compared regarding the surgical results, microscopic structure and inflammatory process exponents of the stomach wall, and receptors for selected tissue growth factors. The IGB group had a longer median hospital stay than that of the non-IGB group. Compared with the non-IGB group, the IGB group had a thicker stomach wall, more submucosal fibrosis, and increased amounts of growth factors and inflammatory markers. Patients with IGB placement before LSG showed greater changes in the stomach wall than those of patients who received LSG alone. IGB placement was associated with stomach muscle layer thickening, submucosal fibrosis, and increased levels of inflammatory markers and tissue growth factors.

CCN2/CTGF-A Modulator of the Optic Nerve Head Astrocyte.

In primary open-angle glaucoma (POAG), a neurodegenerative disease of the optic nerve (ON) and leading cause of blindness, the optic nerve head (ONH) undergoes marked structural extracellular matrix (ECM) changes, which contribute to its permanent deformation and to degeneration of ON axons. The remodeling process of the ECM causes changes in the biomechanical properties of the ONH and the peripapillary sclera, which is accompanied by an increased reactivity of the resident astrocytes. The molecular factors involved in the remodeling process belong to the Transforming growth factor (TGF)-β superfamily, especially TGF-β2. In previous publications we showed that TGF-β2 induced ECM alterations are mediated by Cellular Communication Network Factor (CCN)2/Connective Tissue Growth Factor (CTGF) and recently we showed that CCN2/CTGF is expressed by astrocytes of the ON under normal conditions. In this study we wanted to get a better understanding of the function of CCN2/CTGF under normal and pathologic conditions. To this end, we analyzed the glial lamina and peripapillary sclera of CCN2/CTGF overexpressing mice and studied the effect of CCN2/CTGF and increasing substratum stiffness on murine ON astrocytes in vitro. We observed enhanced astrocyte reactivity in the ONH, increased ECM protein synthesis in the peripapillary sclera and increased Ccn2/Ctgf expression in the ONH during the pathologic development in situ. CCN2/CTGF treatment of primary murine ON astrocytes induced a higher migration rate, and increase of ECM proteins including fibronectin, elastin and collagen type III. Furthermore, the astrocytes responded to stiffer substratum with increased glial fibrillary acidic protein, vimentin, actin and CCN2/CTGF synthesis. Finally, we observed the reinforced appearance of CCN2/CTGF in the lamina cribrosa of glaucomatous patients. We conclude that reactive changes in ONH astrocytes, induced by the altered biomechanical characteristics of the region, give rise to a self-amplifying process that includes increased TGF-β2/CCN2/CTGF signaling and leads to the synthesis of ECM molecules and cytoskeleton proteins, a process that in turn augments the stiffness at the ONH. Such a scenario may finally result in a vicious circle in the pathogenesis of POAG. The transgenic CTGF-overexpressing mouse model might be an optimal model to study the chronic pathological POAG changes in the ONH.