The Inhibitory Effect of Noscapine on the In Vitro Cathepsin G-Induced Collagen Expression in Equine Endometrium

Cathepsin G (CAT) is a protease released by neutrophils when forming neutrophil extracellular traps that was already associated with inducing type I collagen (COL1) in equine endometrium in vitro. Endometrosis is a fibrotic condition mainly characterized by COL1 deposition in the equine endometrium. The objective was to evaluate if noscapine (an alkaloid for cough treatment with anti-neoplastic and anti-fibrotic properties) would reduce COL1A2 transcription (evaluated by qPCR) and COL1 protein relative abundance (evaluated by western blot) induced by CAT in equine endometrial explants from follicular and mid-luteal phases treated for 24 or 48 h. The explants treated with CAT increased COL1 expression. Noscapine decreased COL1A2 transcription at both estrous cycle phases, but COL1 relative protein only at the follicular phase, both induced by CAT. Additionally, the noscapine anti-fibrotic action was found to be more effective in the follicular phase. The CAT treatment caused more fibrosis at the longest period of treatment, while noscapine acted better at the shortest time of treatment. Our results showed that noscapine could act as an anti-fibrotic drug in equine endometrosis by inhibiting CAT in vitro. Noscapine offers a new promising therapeutic tool for treating fibrosis as a single non-selective agent to be considered in the future.

Copaiba oil-resin (Copaifera langsdorfii Desf.: Caesalpiniaceae) associated with laser therapy for skin wound treatment in Wistar rats

Alternative protocols for the treatment of skin lesions have been developed with the use of techniques such as photobiomodulation and phytotherapy, aiming to optimize this process. To evaluate the effectiveness of copaiba (Copaiferalangsdorffii) oil-resin and low-level laser therapy for treating cutaneous wounds, 15 Wistar rats (Rattusnorvergicus) were used, in whom five 8-mm lesions were produced. The following protocols were applied: negative control group (T1); positive control group (T2); laser therapy with AsGa (904 nm), continuous, focal mode for 10 s, dosage of 4 J/cm² (T3); copaiba oil-resin (T4); and association group (copaiba and low-level laser) (T5). The efficacy of each technique was evaluated based on macroscopic aspects of the lesion, wound healing rate, and histopathological analysis (inflammatory infiltrate and collagen expression). The Kruskal-Wallis test was used for statistical analyses (P> 0.05). Copaiba treatment showed an advantage in type III collagen expression, whereas laser therapy demonstrated an enhanced capacity for tissue regeneration. The significant advantage obtained from the association treatment is the improvement of the macroscopic aspect of the wound, with a reduction in crust formation.

The inflammation influence on pathological remodeling of pulmonary arteries in monocrotaline-induced pulmonary hypertension in rats

Aim To investigate the inflammation role on pathological remodeling of pulmonary arteries (PA) in monocrotaline-induced pulmonary hypertension (mPAH) in rats with joint assessment serum and tissue inflammatory biomarkers and the morphological arteries changes. Methods The mPAH was induced by a subcutaneous monocrotaline injection (60 mg/kg) in male rats and control group received a single saline solution. Baseline and every 2, 4, 6, 8 weeks after the serum concentrations of interleukin-6 (IL-6), interleukin-10 (IL-10) were measured by enzyme-linked immunosorbent assay; matrix metalloproteinase-9 (MMP-9), interleukin-1β (IL-1β), collagen type 1 and 3, smooth muscle actin α (SMA-α) in lung tissue were investigated immunohistochemically and quantitative measurements of intima and media thickness were done by planimetry. The functional activity neutrophil changes measured by chemiluminescence and fluorescent methods. Results The IL-10 increased after 2 weeks of mPAH (5,9 vs 0,6 pg/ml, p<0,05) vs control and then it decreased to initial values by 8 weeks (0,06 vs 0,62 pg/ml, p>0,05). The increasing IL-6 (30,3 vs 0,01 pg/ml, p<0,05) and the maximum expression of IL-1β in the lung tissue (0,119 vs 0,099 index of expression (IE), p<0,05) we observed 4 weeks after mPAH. The SMA-α (29,4 vs 40,2 IE, p<0,05) and MMP-9 (1,6 vs 0,8 IE, p<0,05) expression level significantly raised 4 weeks later vs control and remained in a high level until 8 week. A significant increase of type 1 collagen expression was observed at all phases of the experiment, and high level type 3 collagen expression was observed from 4 to 8 weeks (7,0 vs 10,4 IE, p<0,05). The histological characteristics of remodeling these were a thickening of the media (30,6 vs 56,0 μm, p<0,05) and the subintimal layer (1,6 vs 10,9 μm, p<0,05) of the PA. 2 weeks after mPAH cell priming occurred which manifested by modification ROS generation systems, decrease NADPH oxidase activity, increase of myeloperoxidase secretion (MPO), enhance of unbound cytosolic calcium ions, mitochondrial potential reduction. From 4 to 8 weeks an increase NADPH oxidase activity and MPO secretion was revealed into the extracellular environment which leads to overproduction of hypochlorous acid. This functional activity reprogramming of circulating neutrophils indicate associated with time-development of mPAH. Conclusion The inflammation is the most important mediator of pathological remodeling processes in mPA. The monocrotaline launches a neutrophil reaction at an early stage of PAH with changes their functional activity which leads to immune cells recruitment into the lung tissue, producing inflammation and proliferation biomarkers. The hyperplasia of smooth muscle cells and reconstruction of the extracellular matrix are the result of this process and leads to increase intima and media thickness. The high MMP-9, SMA-α, IL-6 activity in 6–8 weeks reflects maintenance local inflammatory potential. FUNDunding Acknowledgement Type of funding sources: Public Institution(s). Main funding source(s): Basic and applied sciences - medicine, subprogram “Diagnostics and therapy of diseases” on the assignment “To establish the molecular-cellular mechanisms of the development of irreversible remodeling of pulmonary vessels in pulmonary arterial hypertension in an experiment in vivo.”

Non-invasive investigation of early kidney damage in streptozotocin-induced diabetic rats by intravoxel incoherent motion diffusion-weighted (IVIM) MRI

Background The current study investigated the performance of intravoxel incoherent motion diffusion (IVIM) technology in monitoring early renal injury in streptozotocin rats. Methods Forty-eight Sprague-Dawley (SD) rats were divided into a control group and a diabetic mellitus (DM) group. Six rats in each group were randomly selected for MR scans at four different time points (0, 4, 8, and 12 weeks). The IVIM-derived parameters (D, D*, f and ADC values) of the renal cortex (CO), outer and inner stripe of the outer medulla (OS, IS), and internal medulla (IM) were acquired. Changes in each IVIM-derived parameter over time were analyzed, and differences between the two groups at each point were assessed. The associations between the IVIM parameters and IV collagen expression, urine volume (UV), blood urea nitrogen (BUN), and serum creatinine (Scr) were investigated. Results The D and D* values of CO and the ADC values of CO, OS, IS and IM displayed significantly different trends between the two groups over time (P<0.05). In addition, significant correlations were discovered between the D* value of CO and UV and BUN (r=0.527, P=0.033; r=0.617, P=0.005), between the ADC value of IM and BUN (r=0.557, P=0.019) and between the f value of IM and BUN (r=0.527, P=0.033). No correlation was found between IVIM parameters and IV collagen expression and Scr. Conclusions IVIM is a potential sensitive and noninvasive technology for the simultaneous assessment of early renal cortical and medullary injuries induced by diabetes.

Low-molecular-weight whey proteins promote collagen production in dermal fibroblasts via the TGF-β receptor/Smad pathway

ABSTRACT Whey proteins (WPs) reportedly enhance cutaneous tissue regeneration in in vivo studies. However, the underlying mechanisms of such regenerative processes are poorly understood. In this study, we show that low-molecular-weight WPs (LMWPs; 1-30 kDa) accelerate the dermal collagen production via the transforming growth factor β receptor (TβR)/Smad pathway. We showed that LMWPs increased type I and III collagen expression in normal human dermal fibroblasts. Moreover, LMWPs rapidly induced Smad protein phosphorylation and nuclear translocation. Notably, type I TβR/Smad signaling inhibitor treatment or type II TβR siRNA knockdown blocked the LMWP-induced type I collagen expression. To identify the active components, we fractionated LMWPs and identified β-lactoglobulin and α-lactalbumin as potential TβR/Smad signaling inducers. Our findings unravel novel biological functions of WPs, involving the TβR/Smad-dependent induction of dermal collagen synthesis, highlighting the therapeutic potential of LMWPs in wound healing.

Preventive training does not interfere with mRNA-encoding myosin and collagen expression during pulmonary arterial hypertension

To gain insight on the impact of preventive exercise during pulmonary arterial hypertension (PAH), we evaluated the gene expression of myosins and gene-encoding proteins associated with the extracellular matrix remodeling of right hypertrophied ventricles. We used 32 male Wistar rats, separated in four groups: Sedentary Control (S, n = 8); Control with Training (T, n = 8); Sedentary with Pulmonary Arterial Hypertension (SPAH, n = 8); and Pulmonary Arterial Hypertension with Training (TPAH, n = 8). All rats underwent a two-week adaptation period; T and TPAH group rats then proceeded to an eight-week training period on a treadmill. At the beginning of the 11th week, S and T groups received an intraperitoneal injection of saline, and SPAH and TPAH groups received an injection of monocrotaline (60 mg/kg). Rats in the T and TPAH groups then continued with the training protocol until the 13th week. We assessed exercise capacity, echocardiography analysis, Fulton’s index, cross-sectional areas of cardiomyocytes, collagen content and types, and fractal dimension (FD). Transcript abundance of myosins and extracellular matrix genes were estimated through reverse transcription-quantitative PCR (RT-qPCR). When compared to the SPAH group, the TPAH group showed increases in functional capacity and pulmonary artery acceleration time/pulmonary ejection time ratio and decreases in Fulton’s index and cross-sectional areas of myocyte cells. However, preventive exercise did not induce alterations in col1a1 and myh7 gene expression. Our findings demonstrate that preventive exercise improved functional capacity, reduced cardiac hypertrophy, and attenuated PH development without interfering in mRNA-encoding myosin and collagen expression during PAH.

The Effect of Topical Corticosteroid Time of Application on Fibroblast and Type III Collagen Expression in Oryctolagus cuniculus with Deep Dermal Burn Wound (As an Indicator for the Best Time to Start Topical Corticosteroid Application in Preventing Hypertrophic Scar)

Background: Hypertrophic scar is an abnormal scar that causes physical deteriorations, psychological problems, and aesthetic issues. An excessive number of fibroblasts and collagen III expressions are histopathology indicators for the hypertrophic scar. The role of topical corticosteroids in suppressing inflammation and hypergranulation had widely demonstrated in previous studies. However, there is no study related to the application of topical corticosteroids as prevention of hypertrophic scars from burn wound found. Hence, this study aimed to examine the evidence of the effects of corticosteroid topical in decreasing the number of fibroblasts and type III collagen expression and the best time to start its application in preventing hypertrophic scars. Methods: This randomized experimental post-test only study involved 54 deep dermal burn wounds on the ventral ear of female Oryctolagus cuniculus that distributed into three groups based on the healing phases. Each group consisted of treatments and controls. Corticosteroid topical application on the first treatment group (inflammatory phase group), the second group (proliferation phase group), and the third group (remodelling phase group) was started on day 3, on day 10, and day 21, respectively. Specimens taken on day 35. Haematoxylin-Eosin and Immunohistochemically staining performed to measure the number of fibroblasts and type III collagen and to observe the epithelization and inflammation process. Results: The number of fibroblasts significantly decreased in the second treatment group (p =0.001) and followed by the first group (p = 0.016), but no significant decrease found in the third group (p = 0.430). The type III collagen decreased significantly in the second treatment group (p = 0.000) and followed by the third group (p = 0.019), but no significant decrease found in the first group. There was no statistically different number of fibroblast and type III collagen discovered between the controls. Complete epithelization found in all groups. Also, no ongoing inflammation found in all groups. Conclusion : Topical corticosteroids on deep dermal burn wound revealed to be effective in reducing the number of fibroblasts and type III collagen with no healing disruption. The proliferation phase found to be the best time to start the application of topical corticosteroids.