Hydroxyproline Content Research Articles

Cloning, tissue distribution and mRNA expression of type I collagen alpha 1 gene from Chu's croaker (Nibea coibor)

The demand for collagen has been increasing over years due to its wide application in food, cosmetics and biomedicine industries. The synthesis of collagen protein in fish depends on instructions provided by collagen, type I, alpha 1 (COL1A1) gene. However, cloning, tissue distribution and mRNA expression of COL1A1 gene in a gel-producing Chu's croaker (Nibea coibor) is currently unknown. This study cloned the cDNA of COL1A1 gene (GenBank accession number: MK641512) from six N. coibor fish. The distribution and mRNA expression pattern of COL1A1 was analyzed in eight tissues of N. coibor. The COL1A1 cDNA had a full length of 6130 bp and contained a 4344 bp open reading frame (ORF) encoding a polypeptide of 1448 amino acids. The homology of N. coibor COL1A1 amino acid had 98% similarity with Larimichthys crocea, indicating conservatism with other members in same family (Sciaenidae). The deduced polypeptide contained the same signal peptides, C-propeptide and N-propeptide domains, and triple helix domains, which are the characteristics of type I collagen in vertebrates. The mRNA of COL1A1 gene was expressed significantly higher in the spine of N. coibor than in all other tissues (P < 0.05), followed by swim bladder, skin and scales. The swim bladder had higher collagen and hydroxyproline contents than other tissues, followed by spine >, scales > and > skin (P < 0.05). Our study successfully cloned the COL1A1 gene from N. coibor for the first time. The COL1A1 gene contained all the features of collagen pro-α1(I) chain proteins, and shared high homology with other marine teleost. COL1A1 gene in N. coibor is highly expressed in spine and swim bladder, consistent with collagen distribution. Our study contributes to better understanding on collagen biosynthesis in N. coibor tissues for various industrial uses.

In vitro and in vivo wound healing activities of Globularia arabica leaf methanolic extract in diabetic rats.

This study assessed the effects of topical application of the plant Globularia arabica extract on the diabetic rate of wound closure. The effect of methanol extracts of G. arabica leaf extract on pro-inflammatory and anti-inflammatory cytokines, as well as the rate of wound contraction, hydroxyproline, and collagen content, was examined using an excision model of wound repair in male rats. The healing pattern was also highly related to using 10% plant extract and significantly, (p-values <0.025) increased wound contraction and increased hydroxyproline and collagen expression in both non-diabetic and diabetic rats, demonstrating that G. arabica extract possesses potent wound healing capacity. During the early wound healing phase, IL-6 levels were found to be upregulated by G. arabica treatment. Increased wound contraction augmented hydroxyproline and collagen content, supporting the early wound healing exhibited by G. arabica. One of the ways for speeding wound healing may be the induction of cytokine production. The result of our study shows that the fibroblast cell line treated with 20µg/ml methanolic extract of G. arabica significantly (p-values <0.035) increases the cell migration that indicated the high wound healing activity. Results suggest that G. arabica may be useful in the topical management of wound healing.

Lycorine Ameliorates Thioacetamide-Induced Hepatic Fibrosis in Rats: Emphasis on Antioxidant, Anti-Inflammatory, and STAT3 Inhibition Effects.

Liver fibrosis is a foremost medical concern worldwide. In Saudi Arabia, numerous risk factors contribute to its high rates. Lycorine—a natural alkaloid—has antioxidant, anti-inflammatory, and antitumor activates. It has been reported to inhibit STAT3 in cancer. Therefore, this study aimed at investigating the possible antifibrotic effect of lycorine against thioacetamide (TAA)-induced liver fibrosis in rats and at elucidating the possible mechanisms. Liver fibrosis was induced by TAA (200 mg/kg i.p.), three per week for four weeks. Treatment with lycorine (0.5 and 1 mg/kg/d) amended TAA-induced rise of serum transaminases that was confirmed histopathologically. Moreover, it ameliorated liver fibrosis in a dose-dependent manner, as indicated by hindering the TAA-induced increase of hepatic hydroxyproline content, α-smooth muscle actin (α-SMA) and transforming growth factor (TGF-β1) expressions. TAA-induced oxidative stress was amended by lycorine treatment via restoring reduced glutathione and diminishing lipid peroxidation. Moreover, lycorine ameliorated hepatic inflammation by preventing the rise of inflammatory cytokines. Notably, lycorine inhibited STAT3 activity, as evidenced by the decreased phospho-STAT3 expression, accompanied by the elevation of the hepatic Bax/Bcl-2 ratio. In conclusion, lycorine hinders TAA-induced liver fibrosis in rats, due to—at least partly—its antioxidative and anti-inflammatory properties, along with its ability to inhibit STAT3 signaling.

In vivo Wound Healing Potential of Raloxifene Nanoemulsion Gel for the Management of Postmenopausal Cutaneous Wounds

Abstract: Background: Depletion in estrogen level(s) especially in postmenopausal women is reported to have delayed wound healing effects; hence we have evaluated the wound healing potential of raloxifene in rat model. Objectives: Investigating the wound healing effects of raloxifene nanoemulsion for the management of postmenopausal cutaneous wounds. Materials and Methods: The optimized nanoemulsion gel contains 0.072% raloxifene hydrochloride. Female Wistar rats were used to investigate its wound healing effects. After three months of ovariectomy, wound healing effect was observed in terms of breaking strength, tensile strength, area of wound contraction, wound closure time, hydroxyproline content and histopathological changes. Results: The nanoemulsion gel exhibited better retention (34.31%) than its nanoemulsion. The raloxifene nanoemulsion gel has no erythema and no eschar formation recorded, and it is safe for topical use. In the incision wound model in ovariectomized rats, breaking (898±25g) and tensile strengths (4.47±0.12 g/mm2) in raloxifene treated groups were found to be higher than the untreated control group. Additionally, in ovariectomized rats, wound contraction was found to be 100% in the treated group s following 20 days of post-wounding, where as in control group only 88% was contraction was observed. Also, more hydroxyproline content in raloxifene treated ovariectomized rat was observed that recommend more collagen content than the untreated ovariectomized rat but approximately similar effects to untreated non-ovariectomized rats. Histopathological studies confirmed that the raloxifene treated groups had more re-epithelialization, neo-vascularization, fibroblast proliferation, and collagen deposition than the control group. Conclusion: These results confirms that the raloxifene nanoemulsion gel has significant wound healing potential, as observed in ovariectomized rats, which will be helpful in postmenopausal cutaneous wound healing. Key words: Raloxifene, Nanoemulsion gel, Ovariectomized, Postmenopausal, Breaking strength, Wound contraction, Hydroxyproline, Histopathology.

Biochemical analysis of collagens from the bone of lizardfish (Saurida tumbil Bloch, 1795) extracted with different acids.

BackgroundLizardfish (Saurida tumbil Bloch, 1795) bone is a fish by-product generated during industrial surimi processing. This by-product is an important source of collagen production since the use of terrestrial animal-based collagens no longer sought due to concern regarding the transfer of infectious diseases and religious issues. Hence, this study was carried out to determine the biochemical analysis of collagens from the bone of lizardfish extracted with different acids.MethodsLizardfish bone collagens were extracted with various acids (i.e., acetic, lactic and citric acids). All extraction processes were conducted in a chiller room (4 °C). The extracted collagens were biochemically characterized, such as hydroxyproline content, Ultraviolet (UV) absorption, X-ray diffraction (XRD), Fourier transform infrared spectroscopy spectra (FTIR), Differential scanning calorimetry (DSC) and solubility in different pH values and NaCl concentrations.ResultsThe yield of extracted collagens ranged between 1.73% and 2.59%, with the highest (p < 0.05) observed in citric acid-extracted collagen (CaEC). Protein patterns confirmed that all-collagen samples had two identical subunits, α1 and α2, representing type I collagen. The highest whiteness value was found in acetic acid-extracted collagen (AaEC), but there was no significant difference (p ≥ 0.05) compared to lactic acid-extracted collagen (LaEC). UV absorption and XRD analysis reflected the characteristics of the collagen, as reported in the literature. For the FTIR, all acid-extracted collagen samples presented a triple helical structure. The thermal transition temperature (Tmax = 77.92–89.04 °C) was in accordance with collagen extracted from other fish species. All extracted collagens were highly soluble in acidic pH and low concentrations of NaCl (0–20 g/L). In conclusion, collagens extracted from lizardfish bone may be used as alternative sources of collagen in industrial settings, and AaEC would be considered superior in terms of the characteristics evaluated in this study.

Preparation of Super Absorbent and Highly Active Fish Collagen Sponge and its Hemostatic Effect in vivo and in vitro.

Effective control of acute wound hemorrhage caused by battlefields, car accidents, natural disasters can highly improve patients’ survival rates. Nevertheless, hemostatic materials on present market have various defects and limitations. This study utilizes tilapia to extract macromolecular type I collagen to prepare a new hemostatic sponge for controlling acute wound bleeding. The extracted fish collagen has high purity, uniform molecular size and high hydroxyproline content. The peptide chain structure and natural high-level structure are intactly preserved. The infrared absorption spectrum showcases that it preserves all the characteristic absorption bands of type I collagen. The developed hemostatic sponge has a uniform honeycomb-shaped porous structure and high water absorption capacity. The biological safety test illustrates that the sponge cell has good compatibility and it will not trigger any inflammatory reaction or immune rejection reactions in the body. The sponge cell could be degraded gradually and completely, which has good biocompatibility and degradation performance. The result of in vitro experiments shows that certain groups or structures in fish collagen molecules can combine specific sites on the surface of blood cells and platelets, which can quickly activate platelets and coagulation system to obtain better coagulation function. The result of In vivo experiments further shows that the fish collagen sponge has fast coagulation speed and low bleeding during the hemostasis process of rabbit ear arteries and rat liver wounds, which proves that it has excellent coagulation performance.

Interactions of dietary eicosapentaenoic acid and vitamin C on growth performance, anti-oxidation and muscle quality of abalone Haliotis discus hannai Ino

A 93-day feeding trial was conducted to investigate the effects of dietary eicosapentaenoic acid (EPA) and vitamin C (VC) on the growth performance, anti-oxidation and muscle quality of abalone Haliotis discus hannai (initial body weight: 26.89 ± 0.26 g; initial shell length: 60.36 ± 0.25 mm). Six experimental diets in a 3 × 2 two-factor design were formulated with graded levels of dietary EPA (0.25%, 0.75% and 1.25%) and dietary VC (0 and 1260 mg/kg). Results showed that the weight gain rate (WGR) of abalone was significantly increased with the increasing dietary EPA levels (P < 0.05), but there was no significant effect of dietary VC on the WGR. The significant highest superoxide dismutase (SOD) activity was observed in the group with 1.25% of dietary EPA and 1260 mg/kg of dietary VC, and there was a significant interaction between dietary EPA and VC (P < 0.05). Dietary EPA supplementation significantly improved muscle hardness and the content of ∑n-3 polyunsaturated fatty acids, volatile compounds, adenosine-5′-monophosphate (AMP) and inosine-5′-monophosphate (IMP) in muscle. Dietary VC supplementation significantly improved the muscle hardness, springiness, chewiness, and the contents of volatile compounds, hydroxyproline and IMP in muscle (P < 0.05). Significant interactions between dietary EPA and VC were observed on volatile compounds, AMP and IMP (P < 0.05). In conclusion, dietary EPA and VC interactively influence the anti-oxidative capability and muscle quality of abalone.

Physicochemical properties of gelatin produced from Nile tilapia skin using chemical and fermentation pretreatments

In this study, we aimed to verify the effectiveness of fermentation as a pretreatment in Nile tilapia (Oreochromis niloticus) skin gelatin production and to assess its effects on the extracted gelatin. Chemical-gelatin (CG) and fermentation-gelatin (FG) were extracted using hot water from tilapia skin after immersion in chemical reagents (sodium hydroxide and butanol) and after fermentation with Bacillus velezensis FEL-BM21, respectively. FG presented a higher recovery (27.40%), yield (49.99%), and imino acid (proline and hydroxyproline) content (177.17 ± 0.35 residues/1000 residues) than CG, but had a lower molecular weight. Fourier transform infrared and circular dichroism spectroscopy confirmed that the collagen was denatured, and scanning electron microscopy revealed that FG had a more continuous filamentous structure than CG. Furthermore, except for transmittance (66.19% at 450 nm and 86.36% at 620 nm), FG exhibited higher gel strength (492.81 g), gelling point (28.93 °C), and melting point (18.33 °C) than CG. These findings suggest that fermentation can be used to treat gelatin raw materials without negatively affecting the structure of gelatin. Moreover, fermentation has potential industrial applications as an alternative to chemical treatment.

Preparation of Codonopsis pilosula polysaccharide microcapsules and its effect and mechanism on skin wound healing in rats.

In this study, after optimizing the extraction process of CPP (Codonopsis pilosula polysaccharides), CPPM (CPP microcapsules) were prepared. Subsequently, the structural characteristics and physicochemical properties were studied. The results showed that CPPM is a hollow sac-like structure with rough folds and protuberances and comes in spherical or ellipsoidal shapes with uniform particle size. CPPM has certain swelling degree, low hardness, good adhesion, and stability. Then, the effect of CPPM on wounds repair was investigated by a rat model. The results showed that CPPM could improve the wound healing rate. Histological evaluation showed CPPM could promote neovascularization and fibroblast proliferation. By investigating the healing mechanism, it was found that CPPM increased the hydroxyproline content in granulation tissue and had an excellent antioxidant ability, and then inhibited lipid peroxidation, in addition, it significantly increased the transcript levels of VEGF and miRNA-21 genes, indicating that CPPM play an influential role in vascular remodeling during wound healing by up-regulating the expression of VEGF and miRNA-21 genes.

Nano-encapsulation of curcumin in fish collagen grafted succinyl chitosan hydrogel accelerates wound healing process in experimental rats

Succinyl chitosan-fish collagen composite hydrogel containing nano-encapsulated curcumin was developed for wound healing application. Curcumin was encapsulated in succinyl chitosan nano-particles using ionic gelation technique. The Z-average diameter of the nanoparticles as observed by dynamic light scattering experiment was ∼47 nm. Similar size ranges were observed in atomic force microscope (AFM) and transmission electron microscopy (TEM) images. A simple one pot gelation of the composite hydrogel containing nano-encapsulated curcumin was achieved by crosslinking succinyl chitosan and fish collagen with caffeic acid. The scanning electron microscopy (SEM) images of the crosslinked hydrogel showed porous sheet like/spongy morphology typical to crosslinked hydrogels. The developed hydrogel was evaluated for its wound healing efficacy in subcutaneous wound model on Wistar albino rats. The hydrogel significantly enhanced the collagen deposition and hydroxyproline content of 599.4 μg/gm and 74.9 μg/gm in wound tissue on the 14th day of post wounding.

Bovine collagen oligopeptides accelerate wound healing by promoting fibroblast migration via PI3K/Akt/mTOR signaling pathway

Bovine collagen oligopeptides (BCOP) are small molecule oligopeptides isolated from bovine collagen, which have been proved to modulate the collagen metabolism. However, the effects of BCOP on wound healing is rarely known by public. This study aims to evaluate the beneficial effects of BCOP on wound healing by using an excisional wound model established in mice. Moreover, the role of the PI3K/Akt/mTOR pathway on the migration of fibroblasts in vitro is further elucidated. It was observed that BCOP could significantly reduce the size of the wound area, increasing albumin and prealbumin levels in mice. BCOP could also inhibit the apoptosis, promoting fibroblast proliferation and migration. The mRNA expressions of Collagen I, which have the ability to transform growth factor-β 1 (TGF-β1) and the hydroxyproline (Hyp) content, were markedly increased after BCOP treatment. The PI3K/Akt/mTOR signaling pathway was also found being activated by BCOP intervention. To summarize, BCOP could accelerate wound healing by promoting fibroblast proliferation, migration and collagen metabolism via PI3K/Akt/mTOR signaling pathway.

Replacement of dietary fish meal with Clostridium autoethanogenum protein on growth performance, digestion, mTOR pathways and muscle quality of abalone Haliotis discus hannai

The present study was conducted to evaluate the replacement of dietary fish meal (FM) with Clostridium autoethanogenum protein (CAP) on abalone Haliotis discus hannai. Five isonitrogenous and isolipidic experimental diets with graded levels of FM replaced by CAP (0, 25, 50, 75 and 100%, respectively) were fed to abalone (initial weight: 21.83 ± 0.25 g) for 110 days. The five diets were named as CAP0, CAP25, CAP50, CAP75 and CAP100, respectively. The results showed that total replacement of dietary FM by CAP did not significantly affect the growth of abalone compared with the control. The highest weight gain rate was observed in the CAP25 group. Compared to the CAP0 group, the CAP25 group also had significantly higher total essential amino acids content in muscle and trypsin activity in digestive gland. The expressions of mammalian target of rapamycin, phosphatidylinositol 3-kinase and threonine kinase/protein B were significantly upregulated, and the expression of AMP-activated protein kinase α2 was significantly downregulated in the muscle of abalone in the CAP25 and CAP50 groups. The crude protein content, hardness, hydroxyproline content, pH value, pyridinium crosslink content, activities of lysyl hydroxylase and prolyl 4-hydroxylase in muscle were significantly elevated in the CAP25 and CAP50 groups. Considering the growth performance, digestion, mTOR pathways and muscle quality, 25% replacement of dietary FM by CAP (4.1% of dietary CAP inclusion level) was recommended for abalone.

Effect of lung pericyte-like cell ablation on the bleomycin model of injury and repair.

We previously showed that pericyte-like cells derived from the FoxD1-lineage contribute to myofibroblasts following bleomycin-induced lung injury. However, their functional significance in lung fibrosis remains unknown. In this study, we used a model of lung pericyte-like cell ablation to test the hypothesis that pericyte-like cell ablation attenuates lung fibrosis in bleomycin-induced lung injury. Lung fibrosis was induced by intratracheal instillation of bleomycin. To ablate pericyte-like cells in the lung, diphtheria toxin (DT) was administered to Foxd1-Cre;Rosa26-iDTR mice at two different phases of bleomycin-induced lung injury. For early ablation, we coadministered bleomycin with DT and harvested mice at days 7 and 21. To test the effect of ablation after acute injury, we delivered DT 7 days after bleomycin administration. We assessed fibrosis by lung hydroxyproline content and semiquantitative analysis of picrosirius red staining. We performed bronchoalveolar lavage to determine cell count and differential. We also interrogated mRNA expression of fibrosis-related genes in whole lung RNA. Compared with DT-insensitive littermates where pericyte-like cells were not ablated, DT-sensitive animals exhibited no difference in fibrosis at day 21 both in the early and late pericyte ablation models. However, early ablation of pericytes reduced acute lung inflammation, as indicated by decreased inflammatory cells. Our data confirm a role for pericytes in regulating pulmonary inflammation in early lung injury.

Revealing potential anti-fibrotic mechanism of Ganxianfang formula based on RNA sequence

BackgroundGanxianfang (GXF) formula as a traditional Chinese medicine (TCM) is used for liver fibrosis in clinical practice while its mechanism is unclear. The aim of this study is to explore the molecular mechanism of GXF against CCl4-induced liver fibrosis rats.MethodsDetected the main compounds of GXF by UPLC-MS/MS. Evaluated the efficacy of GXF (1.58, 3.15, 4.73 g/kg/day) and Fuzheng Huayu (FZHY, positive control, 0.47 g/kg/day) through serum alanine aminotransferase (ALT), aspartate aminotransferase (AST) levels and histopathological changes. Explored the underlying mechanisms by integrating our total liver RNA sequencing (RNA-seq) data with recent liver single-cell sequencing (scRNA-seq) studies. Verified potential pharmacodynamic substances of GXF by hepatic stellate cell (HSC)-T6 line.ResultsMain compounds were identified in GXF by UPLC-MS/MS, including baicalin, wogonoside and matrine etc. With GXF-high dose treatment, the elevation of ALT and AST induced by CCl4 were significantly reduced, and the protective effect of GXF-high dose treatment was better than FZHY. Liver histopathological changes were alleviated by GXF-high dose treatment, the ISHAK scoring showed the incidence of liver cirrhosis (F5/F6) decreased from 76.5 to 55.6%. The results of liver hydroxyproline content were consistent with the histopathological changes. RNA-seq analysis revealed the differential genes (DEGs) were mainly enriched in ECM-receptor interaction and chemokine signaling pathway. GXF effectively inhibited collagen deposition and significantly downregulated CCL2 to inhibit the recruitment of macrophages in liver tissue. Integrating scRNA-seq data revealed that GXF effectively inhibited the expansion of scar-associated Trem2+CD9+ macrophages subpopulation and PDGFRα+PDGFRβ+ scar-producing myofibroblasts in the damaged liver, and remodeled the fibrotic niche via regulation of ligand-receptor interactions including TGFβ/EGFR, PDGFB/PDGFRα, and TNFSF12/TNFRSF12a signaling. In vitro experiments demonstrated that baicalin, matrine and hesperidin in GXF inhibited the activation of hepatic stellate cells.ConclusionsThis study clarified the potential anti-fibrotic effects and molecular mechanism of GXF in CCl4-induced liver fibrosis rats, which deserves further promotion and application.

Properties and Characteristics of Acid-Soluble Collagen from Salmon Skin Defatted with the Aid of Ultrasonication

Salmon skin, a byproduct from the deskinning process, can be used as an alternative source of collagen. Due to the high fat content in skin, the defatting process is required prior to extraction. The properties and characteristics of acid-soluble collagen (ASC) from salmon skin (Oncorhynchus nerka), defatted using isopropanol without and with ultrasonication (70% amplitude for 10 min), were investigated. The ASC from the skin that was defatted with aid of ultrasonication (U-ASC) exhibited lower (p &lt; 0.05) fat content (1.86%) with extraction yield (23.18% w/w, dry weight basis). U-ASC had a higher hydroxyproline content (49.15 mg/g dry sample) and maximum transition temperature (Tmax) (11.6 °C) than C-ASC (collagen extracted from skin defatted without ultrasonication). Both of the ASCs were classified as type I collagen. C-ASC and U-ASC had isoelectric points of 7.17 and 7.40, respectively. Fourier transform infrared (FTIR) and circular dichroism spectra reconfirmed the triple-helix structure of both ASCs. The major amino acid of both collagens was glycine (297–308 residues/1000 residues). A high amount of imino acid (191–193 residues/1000 residues) was also found. After gastrointestinal digestion, the degree of hydrolysis of the digested U-ASC (23.19%) was slightly higher than that of the digested C-ASC (22.31%). However, both digests had no differences in antioxidant activities. Both of the ASCs could be therefore used as functional ingredient.

Improvement of the pharmacokinetics and antifibrotic effects of nintedanib by intrapulmonary administration of a nintedanib–hydroxypropyl-γ-cyclodextrin inclusion complex in mice with bleomycin-induced pulmonary fibrosis

Idiopathic pulmonary fibrosis is a chronic lung disease that is characterized by progressive abnormal reprogramming following injury of the pulmonary structure. In this study, we prepared a nintedanib (antifibrotic agent) and cyclodextrin (CyD) inclusion complex to improve the pharmacokinetics and antifibrotic effects of nintedanib following intrapulmonary administration. Hydroxypropyl-γ-CyD (HP-γ-CyD) enhanced the solubility of nintedanib without cytotoxic effects on WI-38 cells (lung fibroblasts) and NCI-H441 cells (alveolar epithelium model). Compared with nintedanib ethanesulfonate salt, the nintedanib–HP-γ-CyD inclusion complex exhibited prolonged distribution in the lungs following intrapulmonary administration in mice with bleomycin-induced pulmonary fibrosis. In addition, compared with nintedanib ethanesulfonate salt, the nintedanib–HP-γ-CyD inclusion complex exhibited higher stability in the bronchoalveolar lavage fluid and lower permeability in NCI-H441 cell monolayers. These results suggested that the inclusion complexation of nintedanib into HP-γ-CyD improved its pharmacokinetics following intrapulmonary administration by increasing its stability in the lungs and reducing its permeability through the alveolar cell membrane. Intrapulmonary administration of the nintedanib–HP-γ-CyD inclusion complex significantly reduced the intrapulmonary hydroxyproline content and limited pathological fibrotic changes. Overall, this study indicates that antifibrotic agent–CyD inclusion complexation intended for intrapulmonary administration can be used to prolong distribution in the lungs and lead to the expansion of idiopathic pulmonary fibrosis therapy.

Nifuroxazide ameliorates pulmonary fibrosis by blocking myofibroblast genesis: a drug repurposing study

BackgroundIdiopathic pulmonary fibrosis (IPF) is a serious interstitial lung disease with a complex pathogenesis and high mortality. The development of new drugs is time-consuming and laborious; therefore, research on the new use of old drugs can save time and clinical costs and even avoid serious side effects. Nifuroxazide (NIF) was originally used to treat diarrhoea, but more recently, it has been found to have additional pharmacological effects, such as anti-tumour effects and inhibition of inflammatory diseases related to diabetic nephropathy. However, there are no reports regarding its role in pulmonary fibrosis.MethodsThe therapeutic effect of NIF on pulmonary fibrosis in vivo was measured by ELISA, hydroxyproline content, H&E and Masson staining, immunohistochemistry (IHC) and western blot. Immune cell content in lung tissue was also analysed by flow cytometry. NIF cytotoxicity was evaluated in NIH/3T3 cells, human pulmonary fibroblasts (HPFs), A549 cells and rat primary lung fibroblasts (RPLFs) using the MTT assay. Finally, an in vitro cell model created by transforming growth factor-β1 (TGF-β1) stimulation was assessed using different experiments (immunofluorescence, western blot and wound migration assay) to evaluate the effects of NIF on the activation of NIH/3T3 and HPF cells and the epithelial-mesenchymal transition (EMT) and migration of A549 cells.ResultsIn vivo, intraperitoneal injection of NIF relieved and reversed pulmonary fibrosis caused by bleomycin (BLM) bronchial instillation. In addition, NIF inhibited the expression of a variety of cellular inflammatory factors and immune cells. Furthermore, NIF suppressed the activation of fibroblasts and EMT of epithelial cells induced by TGF-β1. Most importantly, we used an analytical docking experiment and thermal shift assay to further verify that NIF functions in conjunction with signal transducer and activator of transcription 3 (Stat3). Moreover, NIF inhibited the TGF-β/Smad pathway in vitro and decreased the expression of phosphorylated Stat3 in vitro and in vivo.ConclusionTaken together, we conclude that NIF inhibits and reverses pulmonary fibrosis, and these results support NIF as a viable therapeutic option for IPF treatment.Graphic

Pharmacological Mechanisms of Shangke Huangshui against Skin and Soft Tissue Infection.

Background Skin and soft tissue infections (SSTIs) are a group of common diseases, usually caused by bacteria. Shangke Huangshui (SK) has been widely used to treat various SSTIs diseases for many years, but its mechanism is unclear. Therefore, this study was designed to investigate the anti-infective effect of SK on different skin and soft tissue infection diseases and to explore its underlying mechanism. Methods The subcutaneous abscess mouse model, the dermal ulcer rat model, and the infectious soft tissue injury rat model were established by injection of Staphylococcus aureus to evaluate the anti-inflammatory and antibacterial effects of SK. Abscess volume, local appearance score and histological changes, bacterial contents, and hydroxyproline concentration in the skin or soft tissue were analyzed. The levels of NO, TNF-α, IL-1β, and IL-8 in the serum and tissue were determined by ELISA method. The mRNA expression levels of TLR2, MyD88, TAK1, NF-κB, AP-1, and other genes were measured with qRT-PCR method, and the protein expression of TLR2, MyD88, TAK1, NF-κB, and AP-1 was detected by western blot method. Results SK had an obvious therapeutic effect on skin and soft tissue infections. It reduced the volume of abscess and promoted the healing of skin ulcer, improved pathological phenomena, such as inflammatory infiltration of skin and soft tissue, reduced the levels of white blood cells and NO in the blood, decreased bacteria contents in the skin and soft tissue. Furthermore, SK decreased the mRNA expression of TLR2, MyD88, TAK1, NF-κB and AP-1, and other related genes and also downregulated the protein expression of TLR2, MyD88, TAK1, NF-κB, and AP-1. Conclusion The experiments provide evidence that SK can treat skin and soft tissue infection diseases based on its comprehensive antibacterial and anti-inflammatory effects. The therapeutic mechanism may be associated with the inhibition of TLR2/MyD88/NF-κB signaling pathway.

Kinetic evaluation of gelatin extraction from chicken skin and the effect of some extraction parameters

AbstractKinetic models have been extensively studied for the prediction of yield and quality for plant‐sourced ingredients although that is very rare for animal‐sourced products. This study is designed to evaluate common kinetic models for the prediction of yield in gelatin extraction from chicken skin. Kinetic models used for comparison were the Peleg's, the first order, and the rate law models while extraction parameters studied were solvent pH, solid:liquid ratio, and sample size. Extraction is evaluated based on the protein and hydroxyproline (Hyp) concentrations of each extract. The success of the models is evaluated based on the root‐mean‐square error (RMSE) and the coefficient of determination (R2) values calculated for fittings of the models to the experimental data. The protein content of the dried and degreased skin was over 50% before gelatin extraction. Neutral pH led to the highest yield for both protein and Hyp, which were close to 2,100 and 200 mg per 10 g of skin, respectively. Similarly, smaller sample size or lower solid:liquid ratio resulted in a higher yield. In general, the first 60 min of the extraction was by rapid washing and later on by slow diffusion. Peleg's model is found to be the best fit to the experimental data with the highest R2 values between 0.97 and 0.99 and the lowest RMSE values compared to that of other kinetic models. This is the first report on the kinetics of gelatin extraction as well as one of the few studies on extraction kinetics of animal‐sourced products.Practical ApplicationsExtraction parameters including the size of the raw material and raw material:solvent ratio showed significant effects on extraction yield. The smaller the size and the ratio, the higher the yield. Kinetics of gelatin extraction is reported for the first time by this manuscript, dealing with the yield during the whole extraction process. Extraction parameters may create substantial variability in quality and yield; thus, the data presented here may be utilized in minimizing those potential variations, by minimizing the time, energy and solvent required. The gelatin industry may use this report in the design of the gelatin extraction process as the data presented concludes that a high‐rate extraction was completed within the first hour of the extraction, leading over 80% of the expected yield. Gelatin manufacturers and researchers in this field may find this manuscript useful as it provides valuable insights on gelatin extraction and its kinetics.

Kidney and lung injury in rats following acute diquat exposure.

Diquat (1,1'-ethylene-2,2'-bipyridylium) is a type of widely used agricultural chemical, whose toxicity results in damage to numerous tissues, including the lung, liver, kidney and brain. The aim of the present study was to establish a rat model of acute diquat exposure and explore the relationship between diquat concentration, and kidney and lung injury, in order to provide an experimental basis for clinical treatment. A total of 140 healthy adult male Wistar rats were randomly divided into control and exposure groups. The diquat solution was administered intragastrically to the exposure group at 1/2 of the lethal dose (140 mg/kg). An equal volume of water was administered to the control group. The dynamic changes in the plasma and tissue diquat levels were quantitatively determined at 0.5, 1, 2, 4, 8, 16 and 24 h following exposure using liquid chromatography mass spectrometry. The content of hydroxyproline (HYP) in the lung tissues, as well as the levels of blood urea nitrogen (BUN), creatinine (Cr), uric acid (UA), kidney injury molecule-1 (KIM-1) and tumor growth factor (TGF)-β1, were detected using western blot analysis at every time point. Lung and kidney morphology were also assessed. Electron microscopy showed that the degree of renal damage gradually increased with time. Vacuolation gradually increased, some mitochondrial bilayer membrane structures disappeared and lysosomes increased. The lung tissue damage was mild, and the cell membrane integrity and organelles were damaged to varying degrees. The plasma and organ levels of diquat peaked at ~2 h, followed by a steady decrease, depending on the excretion rate. Over time, the serum concentrations of UA, BUN, Cr and KIM-1 were all significantly increased (P<0.05). Serum KIM-1 in rats was increased after 0.5 h, and was significantly increased after 4 h, suggesting that KIM-1 is an effective predictor of early renal injury. Early TGF-β1 expression was clearly observed in renal tissue, while no clear TGF-β1 expression was observed in the lung tissue. In conclusion, the concentration of diquat in the serum and tissue of rats with acute diquat poisoning peaked at an early stage and then rapidly decreased. The renal function damage and pathological changes persisted, the lung tissue was slightly damaged with inflammatory cell infiltration, and early pulmonary fibrosis injury was not obvious.