Recombinant collagen coating 3D printed PEGDA hydrogel tube loading with differentiable BMSCs to repair bile duct injury.

Bile duct injury presents a significant clinical challenge following hepatobiliary surgery, necessitating advancements in the repair of damaged bile ducts is a persistent issue in biliary surgery. 3D printed tubular scaffolds have emerged as a promising approach for the repair of ductal tissues, yet the development of scaffolds that balance exceptional mechanical properties with biocompatibility remains an ongoing challenge. This study introduces a novel, bio-fabricated bilayer bile duct scaffold using a 3D printing technique. The scaffold comprises an inner layer of polyethylene glycol diacrylate (PEGDA) to provide high mechanical strength, and an outer layer of biocompatible, methacryloylated recombinant collagen type III (rColMA) loaded with basic fibroblast growth factor (bFGF)-encapsulated liposomes (bFGF@Lip). This design enables the controlled release of bFGF, creating an optimal environment for the growth and differentiation of bone marrow mesenchymal stem cells (BMSCs) into cholangiocyte-like cells. These cells are instrumental in the regeneration of bile duct tissues, evidenced by the pronounced expression of cholangiocyte differentiation markers CK19 and CFTR. The PEGDA//rColMA/bFGF@Lip bilayer bile duct scaffold can well simulate the bile duct structure, and the outer rColMA/bFGF@Lip hydrogel can well promote the growth and differentiation of BMSCs into bile duct epithelial cells. In vivo experiments showed that the scaffold did not cause cholestasis in the body. This new in vitro pre-differentiated active 3D printed scaffold provides new ideas for the study of bile duct tissue replacement.

Morphological adaptation of the tongue of okapi (Okapia johnstoni Artiodactyla, Giraffidae)-Anatomy, histology, and ultrastructure.

The aim of this study was to describe the morphology of the tongue of the okapi, and to compare the results with other ruminants including browsers, intermediates and grazers. The material was collected post-mortem from two animals from a Zoological Garden. The structure of the okapi tongue, focusing of the shape of the tongue, lingual surface, its papillae and lingual glands, was examined using gross morphology, light and polarized microscopy, and by scanning electron microscopy. The okapi tongue was characterized by dark pigmentation on the lingual dorsum (except lingual torus) and on the whole ventral surface. Two types of filiform papillae were observed, with additional, even 6-8 projections at their base. The round fungiform papillae were present at a higher density, up to 16/cm2, on the ventro-lateral area of the lingual apex. Round and elongate vallate papillae were arranged in two parallel lines between the body and root of the tongue. Numerous taste buds were detected within the epithelium of their vallum, while fungiform papillae had sparse taste buds. A lack of foliate papillae was noted. Very small conical papillae, some lenticular in shape, were present on the lingual torus. Thick collagen type I fibers were dominant over collagen type III fibers in the connective tissue of the lingual papillae. The mucous acini units were dominant among lingual glands, indicating that the secretion of okapi lingual glands was mostly mucous. In many aspects, the tongue of okapi resembles the tongue of other ruminants. The specific lingual shape and lingual surface, together with the lingual glands, support the processing of plant food, such as young and soft leaves. Although okapi tongue is characterized by smaller conical papillae compared to other ruminants, its high number of vallate papillae is similar that found in other browsers, intermediate and grazers. Thus the number of gustatory papillae rather indicates that this feature is not related to the type of feeding.

An endometrial biomimetic extracellular matrix (ECM) for enhanced endometrial regeneration using hyaluronic acid hydrogel containing recombinant human type III collagen.

Endometrial injury poses a significant challenge in tissue regeneration, with type III collagen (COL III) playing a pivotal role in maintaining endometrial integrity and facilitating repair. Our study explored the utility of recombinant human type III collagen (RHC) as an intervention for endometrial damage. To address the challenges associated with the inherent instability and rapid degradation of COL III in vivo, we developed an RHC-HA hydrogel by conjugating RHC with hyaluronic acid (HA), thus ensuring a more stable and sustained delivery. Our findings suggested that the RHC-HA hydrogel significantly promoted endometrial regeneration and restored fertility. The hydrogel facilitated prolonged retention of RHC in the uterus, leading to a substantial improvement in the repair process. The synergistic interaction between RHC and HA greatly enhances cell proliferation and adhesion, surpassing the efficacy of HA or RHC alone. Additionally, the RHC-HA hydrogel demonstrated notable anti-fibrotic effects, which are crucial for preventing abnormalities during endometrial healing. These findings suggested that the RHC-HA hydrogel presented a therapeutic strategy in the treatment of uterine endometrial injuries, which may improve female reproductive health.

Recombinant Human Collagen Type III Improves Hypertrophic Scarring by Regulating the Ratio of Type I/III Collagen.

Hypertrophic scar development is a complication associated with wound healing, impacting local appearance and function. The type I/III collagen ratio affects the extent of hypertrophic scarring; a reduced ratio can ameliorate this. In this study, recombinant human collagen type III was developed. Liquid chromatography-tandem mass spectrometry was used to determine its amino acid sequence and confirm its high level of homology with natural human type III collagen. Recombinant human collagen type III displayed no cytotoxicity and did not confer skin irritation and sensitization. Immunofluorescence and western blot analyses of histidine following incubation with fibroblasts suggested cell entry of recombinant human collagen type III. Furthermore, recombinant human collagen type III promoted the synthesis of the natural type III collagen in fibroblasts, resulting in a more obvious increase of type III collagen content in fibroblasts than that of type I collagen, and then decreased the ratio of type I/III collagen. The results of 5-ethynyl-2'-deoxyuridine staining assay suggested enhanced fibroblast proliferation. Following local injection of recombinant human collagen type III, rabbit ear scarring was significantly reduced after 60 days. Vancouver Scar Scale evaluation showed that all index scores were significantly reduced. Western blotting and Picro-Sirius red staining showed that the natural type III collagen increase in scar tissue was greater than that of type I collagen, decreasing the type I/III ratio. In summary, recombinant human collagen type III can be taken up by fibroblasts and promote natural collagen synthesis-especially that of type III-thereby reducing the type I/III ratio and improving hypertrophic scarring.

Efficacy and Safety of Type III Collagen Lyophilized Fibers Using Mid-to-Deep Dermal Facial Injections for the Correction of Dynamic Facial Wrinkles.

BACKGROUND: This study aimed to evaluate the therapeutic efficacy and safety of injecting Type III collagen lyophilized fibers into the mid-to-deep layers of the facial dermis to ameliorate dynamic facial wrinkles. METHODS: In this retrospective analysis, clinical data were collected from patients exhibiting dynamic facial wrinkles (encompassing frown lines, forehead lines, and crow's feet) with a wrinkle severity rating scale (WSRS) score of 3 or higher. In the control group, 75 participants received collagen implant injections into the mid-to-deep facial dermal layers, whereas 76 participants in the experimental group received injections of Type III collagen lyophilized fibers in similar layers. The study analyzed and compared clinical efficacy, WSRS score alterations, patient satisfaction, and safety profiles between the groups over the 30-day and 90-day treatment periods. RESULTS: At the 30-day mark, the therapeutic efficacy was not significantly different between the two groups (P > 0.05). However, at 90 days, the treatment efficacy in the experimental group surpassed that in the control group, showing a statistically significant difference (P < 0.05). After 30 days of treatment, the WSRS score improvement in the experimental group was significantly superior to that in the control group (P < 0.05). Conversely, at the 90-day mark, the results revealed no significant variation in WSRS score improvements between the two groups (P > 0.05). Regarding treatment satisfaction among researchers and participants post-30 and 90-day treatment in both groups, no statistically significant differences were observed (P > 0.05). Similarly, the incidence of adverse reactions between the groups was not statistically significant (P > 0.05). CONCLUSIONS: Injections of lyophilized type III collagen fibers into the mid-to-deep layers of the facial dermis have a definitive therapeutic effect on dynamic facial wrinkles. This treatment not only substantially diminishes wrinkle severity but also has a commendable safety profile. LEVEL OF EVIDENCE I: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Study on the Effect of Type III Recombinant Humanized Collagen on Human Vascular Endothelial Cells.

The effect and mechanism of type III recombinant humanized collagen (hCOLIII) on human vascular endothelial EA.hy926 cells at the cellular and molecular levels were investigated. The impact of hCOLIII on the proliferation of EA.hy926 cells was detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromid assay, the effect of hCOLIII on cell migration was investigated by scratch assay, the impact of hCOLIII on cell cycle and apoptosis was detected by flow cytometry, the ability of hCOLIII to induce angiogenesis of EA.hy926 cells was evaluated by angiogenesis assay, and the effect of hCOLIII on vascular endothelial growth factor (VEGF) expression was detected by real-time reverse transcription-polymerase chain reaction analysis. The hCOLIII at concentrations of 0.5, 0.25, and 0.125 mg/mL all showed specific effects on the proliferation and migration of human vascular endothelial cells. It could also affect the cell cycle, increase the proliferation index, and increase the expression level of VEGF in human vascular endothelial cells. In the meantime, hCOLIII at the concentration of 0.5 mg/mL also showed a promoting effect on vessel formation. hCOLIII can potentially promote the endothelization process of blood vessels, mainly by affecting the proliferation, migration, and vascular-like structure of human endothelial cells. At the same time, hCOLIII can promote the expression of VEGF. This collagen demonstrated its potential as a raw material for cardiovascular implants.

Sodium alginate hydrogel integrated with type III collagen and mesenchymal stem cell to promote endometrium regeneration and fertility restoration.

A thinner endometrium has been linked to implantation failure, and various therapeutic strategies have been attempted to improve endometrial regeneration, including the use of mesenchymal stem cells (MSCs). However, low survival and retention rates of transplanted stem cells are main obstacles to efficient stem cell therapy in thin endometrium. Collagen type III is a key component of the extracellular matrix, plays a crucial role in promoting cell proliferation and differentiation, and has been identified as the major collagen expressed at the implantation site. Herein, composite alginate hydrogel containing recombinant type III collagen (rCo III) and umbilical cord mesenchymal stem cells are developed. rCo III serves as favorable bioactive molecule, displaying that rCo III administration promotes MSCs proliferation, stemness maintenance and migration. Moreover, rCo III administration enhances cell viability and migration of mouse endometrial stromal cells (ESCs). In a mouse model of thin endometrium, the Alg-rCo III hydrogel loaded with MSCs (MSC/Alg-rCo III) significantly induces endometrial regeneration and fertility enhancement in vivo. Further studies demonstrate that the MSC/Alg-rCo III hydrogel promoted endometrial function recovery partly by regulating mesenchymal-epithelial transition of ESCs. Taken together, the combination of Alg-rCo III hydrogel and MSCs has shown promising results in promoting endometrium regeneration and fertility restoration, and may provide new therapeutic options for endometrial disease.

A Rare Case of Diabetic Nephropathy and Type III Collagen Glomerulopathy.

Collagenofibrotic glomerulopathy (CG) is a poorly understood kidney disease characterized by extensive deposition of abnormal type III collagen within the glomeruli. We report the case of a 25-year-old man with known type I diabetes mellitus who presented to the emergency department with diaphoresis, nephrotic-range proteinuria, hypertension, and elevated creatinine. Renal biopsy revealed combined collagenofibrotic glomerulopathy and diabetic nephropathy. This case highlights the clinicopathological features of type III collagen glomerulopathy and its possible association with diabetic nephropathy.

Histopathological Study on Collagen in Full-Thickness Wound Healing in Fraser's Dolphins (Lagenodelphis hosei).

Fraser's dolphins (Lagenodelphis hosei) possess great healing abilities. Their skin composition can be restored after wounding, including collagen spacing, orientation, and bundle thickness. However, it remains unclear how collagens are involved in the wound-healing process and eventually regain normality in Fraser's dolphins. Learned from the other two scarless healing animals, changes in type III/I collagen composition are believed to modulate the wound healing process and influence the scarring or scarless fate determination in human fetal skin and spiny mouse skin. In the current study, Herovici's, trichrome, and immunofluorescence staining were used on normal and wounded skin samples in Fraser's dolphins. The results suggested that type I collagens were the main type of collagens in the normal skin of Fraser's dolphins, while type III collagens were barely seen. During the wound healing process, type III collagens showed at early wound healing stages, and type I collagen increased in the mature healed wound. In an early healed wound, collagens were organized in a parallel manner, showing a transient hypertrophic-like scar, and eventually restored to normal collagen configuration and adipocyte distribution in the mature healed wound. The remarkable ability to remove excessive collagens merits further investigation to provide new insights into clinical wound management.

Recombinant humanized collagen remodels endometrial immune microenvironment of chronic endometritis through macrophage immunomodulation.

Recently, evidence has suggested that chronic endometritis (CE) is a crucial factor associated with infertility and failure of assisted reproductive techniques, prompting concern in the reproductive field. Studies have shown that persistent infiltered immune cells stimulation result in the disturbance of endometrial immune microenvironment could lead to the infertility of CE patients finally. Conventional treatments are limited because they lack immune regulation, so it is urgent to develop a novel approach to treat CE and promote embryo implantation in patients with CE. Herein, we prepared recombinant humanized type III collagen (rhCol III) with high cell adhesion activity to regulate macrophages and repair the endometrium. In this study, M1 macrophages and M1 macrophages cultured medium and lipopolysaccharide (LPS) co-stimulated inflammatory endometrium stromal cells (ESCs) were established in vitro to mimic CE condition. rhCol III promoted M1 macrophages toward M2 phenotype, improved cell migration, viability and collagen components of inflammatory ESCs. Also, the inflammatory response of inflammatory ESCs was downregulated after rhCol III treatment. Subsequently, LPS was used for CE rat model and a 28-day observation was performed; inflammatory cells' infiltration, endometrium repair, extracellular matrix (ECM) remodeling and pregnancy outcomes were promoted after rhCol III endometrial infusion. In conclusion, rhCol III promoted (i) macrophage polarization toward M2 macrophages, (ii) pro-inflammatory cytokine production and anti-inflammatory cytokine reduction, (iii) ECM remodeling and (iv) fertility restoration. Meanwhile, rhCol III enhanced cell biological functions by interacting with discoidin domain receptors, regulated cell metabolism and reduced the inflammatory response through the inhibition of the NF-κB/YAP signaling pathway. Overall, the results illustrated the potential therapeutic prospects of rhCol III for CE treatment.

An experimental study of the vascular embolism caused by recombinant type III collagen implants and hyaluronic acid.

BACKGROUND: There are significant differences in the reported incidence of vascular complications that result from the injection of different soft tissue fillers. This study aimed to compare the risk of vascular embolism after recombinant type III collagen implants and hyaluronic acid (HA) injection into arteries. METHODS: Different concentrations of recombinant type III collagen and Restylane were injected into the central ear artery of rabbits, to construct an immediate embolization model. We screened for vascular recanalization and tissue necrosis at 30 min, 1 day, and 7 days after injection, and histopathology examination was processed on Day 7. RESULTS: At 30 min after injection, complete recanalization of the central ear artery was observed in 17 rabbits in the C1 group while none in the HA group. On Day 1 after injection, complete recanalization of the CEA main trunk was observed in all rabbits in the collagen group while 50% in the HA group. There was a significant difference between the C1 group and the HA group in terms of vascular recanalization and skin necrosis. CONCLUSION: Under the present experimental conditions, the risk of causing vascular embolism was much lower with collagen than with Restylane. Different doses of collagen at different injection rates have the same safety profile.

Effect and mechanism of signal peptide and maltose on recombinant type III collagen production in Pichia pastoris.

Recombinant type III collagen plays an important role in cosmetics, wound healing, and tissue engineering. Thus, increasing its production is necessary. After an initial increase in output by modifying the signal peptide, we showed that adding 1% maltose directly to the medium increased the yield and reduced the degradation of recombinant type III collagen. We initially verified that Pichia pastoris GS115 can metabolize and utilize maltose. Interestingly, maltose metabolism-associated proteins in Pichia pastoris GS115 have not yet been identified. RNA sequencing and transmission electron microscopy were performed to clarify the specific mechanism of maltose influence. The results showed that maltose significantly improved the metabolism of methanol, thiamine, riboflavin, arginine, and proline. After adding maltose, the cell microstructures tended more toward the normal. Adding maltose also contributed to yeast homeostasis and methanol tolerance. Finally, adding maltose resulted in the downregulation of aspartic protease YPS1 and a decrease in yeast mortality, thereby slowing down recombinant type III collagen degradation. KEY POINTS: • Co-feeding of maltose improves recombinant type III collagen production. • Maltose incorporation enhances methanol metabolism and antioxidant capacity. • Maltose addition contributes to Pichia pastoris GS115 homeostasis.

The Associations of Single Nucleotide Polymorphisms of the COL3A1, COL6A5, and COL8A1 Genes with Atopic Dermatitis.

The pathophysiology of atopic dermatitis (AD) is complex, multifactorial, and not fully understood. Genes encoding collagens, the most abundant proteins in the extracellular matrix (ECM), may play a potential role in the pathogenesis of AD. Our study aimed to estimate the associations between Col3A1/rs1800255, Col6A5 /29rs12488457, and Col8A1/rs13081855 polymorphisms and the occurrence, course, and features of AD in the Polish population. Blood samples were collected from 157 patients with AD and 111 healthy volunteers. The genotype distribution of the investigated collagens genes did not differ significantly between the AD and control subjects (p > 0.05). The AA genotype of Col3A1/rs1800255 was significantly associated with the occurrence of mild SCORAD (OR = 0.16; 95% Cl: 0.03-0.78; p = 0.02) and mild pruritus (OR = 18.5; 95% Cl: 3.48-98.40; p = 0.0006), while the GG genotype was significantly associated with severe SCORAD (OR = 6.6; 95% Cl: 1.23-32.35; p = 0.03). Regarding Col6A5/29rs12488457 polymorphism, the average SCORAD score was significantly lower in the group of patients with genotype AA than in patients with the AC genotype (39.8 vs. 53.4; p = 0.04). Nevertheless, both average SCORAD scores were high, and represent the moderate and severe grades of the diseases, respectively. The single nucleotide polymorphisms (SNPs) of COL3A1/ rs1800255 and Col6A5/29rs12488457 seem to be associated with AD courses and symptoms, suggesting new disease biomarkers. The modulation of collagens, the major component of the ECM, may serve as a therapeutic target of AD in the future.

Tumor-restrictive type III collagen in the breast cancer microenvironment: prognostic and therapeutic implications.

Collagen plays a critical role in regulating breast cancer progression and therapeutic resistance. An improved understanding of both the features and drivers of tumor-permissive and -restrictive collagen matrices are critical to improve prognostication and develop more effective therapeutic strategies. In this study, using a combination of in vitro, in vivo and in silico experiments, we show that type III collagen (Col3) plays a tumor-restrictive role in human breast cancer. We demonstrate that Col3-deficient, human fibroblasts produce tumor-permissive collagen matrices that drive cell proliferation and suppress apoptosis in noninvasive and invasive breast cancer cell lines. In human TNBC biopsy samples, we demonstrate elevated deposition of Col3 relative to type I collagen (Col1) in noninvasive compared to invasive regions. Similarly, in silico analyses of over 1000 breast cancer patient biopsies from The Cancer Genome Atlas BRCA cohort revealed that patients with higher Col3:Col1 bulk tumor expression had improved overall, disease-free and progression-free survival relative to those with higher Col1:Col3 expression. Using an established 3D culture model, we show that Col3 increases spheroid formation and induces formation of lumen-like structures that resemble non-neoplastic mammary acini. Finally, our in vivo study shows co-injection of murine breast cancer cells (4T1) with rhCol3-supplemented hydrogels limits tumor growth and decreases pulmonary metastatic burden compared to controls. Taken together, these data collectively support a tumor-suppressive role for Col3 in human breast cancer and suggest that strategies that increase Col3 may provide a safe and effective modality to limit recurrence in breast cancer patients.

Effect of pegbelfermin on NASH and fibrosis-related biomarkers and correlation with histological response in the FALCON 1 trial.

Background & Aims: FALCON 1 was a phase IIb study of pegbelfermin in patients with non-alcoholic steatohepatitis (NASH) and stage 3 fibrosis. This FALCON 1 post hoc analysis aimed to further assess the effect of pegbelfermin on NASH-related biomarkers, correlations between histological assessments and non-invasive biomarkers, and concordance between the week 24 histologically assessed primary endpoint response and biomarkers. Methods: Blood-based composite fibrosis scores, blood-based biomarkers, and imaging biomarkers were evaluated for patients with available data from FALCON 1 at baseline through week 24. SomaSignal tests assessed protein signatures of NASH steatosis, inflammation, ballooning, and fibrosis in blood. Linear mixed-effect models were fit for each biomarker. Correlations and concordance were assessed between blood-based biomarkers, imaging, and histological metrics. Results: At week 24, pegbelfermin significantly improved blood-based composite fibrosis scores (ELF, FIB-4, APRI), fibrogenesis biomarkers (PRO-C3 and PC3X), adiponectin, CK-18, hepatic fat fraction measured by MRI-proton density fat fraction, and all four SomaSignal NASH component tests. Correlation analyses between histological and non-invasive measures identified four main categories: steatosis/metabolism, tissue injury, fibrosis, and biopsy-based metrics. Concordant and discordant effects of pegbelfermin on the primary endpoint vs. biomarker responses were observed; the most clear and concordant effects were on measures of liver steatosis and metabolism. A significant association between hepatic fat measured histologically and by imaging was observed in pegbelfermin arms. Conclusions: Pegbelfermin improved NASH-related biomarkers most consistently through improvement of liver steatosis, though biomarkers of tissue injury/inflammation and fibrosis were also improved. Concordance analysis shows that non-invasive assessments of NASH support and exceed the improvements detected by liver biopsy, suggesting that greater consideration should be given to the totality of available data when evaluating the efficacy of NASH therapeutics. Clinical trial number: Post hoc analysis of NCT03486899. Impact and implications: FALCON 1 was a study of pegbelfermin vs. placebo in patients with non-alcoholic steatohepatitis (NASH) without cirrhosis; in this study, patients who responded to pegbelfermin treatment were identified through examination of liver fibrosis in tissue samples collected through biopsy. In the current analysis, non-invasive blood- and imaging-based measures of fibrosis, liver fat, and liver injury were used to determine pegbelfermin treatment response to see how they compared with the biopsy-based results. We found that many of the non-invasive tests, particularly those that measured liver fat, identified patients who responded to pegbelfermin treatment, consistent with the liver biopsy findings. These results suggest that there may be additional value in using data from non-invasive tests, along with liver biopsy, to evaluate how well patients with NASH respond to treatment.

Recombination humanized type III collagen promotes oral ulcer healing.

OBJECTIVE: Recombinant humanized type III collagen (rhCol III) is a highly adhesive biomaterial composed of 16 adhesion-related tandem repeats refined from human type III collagen. Here, we aimed to investigate the effect of rhCol III on oral ulcers and reveal the underlying mechanism. METHODS: Acid-induced oral ulcers were induced on the murine tongue, and rhCol III or saline drops were administered. The effect of rhCol III on oral ulcers was assessed using gross and histological analyses. The effects on the proliferation, migration, and adhesion of human oral keratinocytes were investigated in vitro. The underlying mechanism was explored using RNA sequencing. RESULTS: Administration of rhCol III accelerated the lesion closure of oral ulcers, reduced the release of inflammatory factors, and alleviated pain. rhCol III promoted the proliferation, migration, and adhesion of human oral keratinocytes in vitro. Mechanistically, the enrichment of genes associated with the Notch signaling pathway was upregulated after rhCol III treatment. CONCLUSION: rhCol III promoted the healing of oral ulcers, showing promising therapeutic potential in oral clinics.

A randomized, double-blind, placebo-controlled phase IIa trial of efruxifermin for patients with compensated NASH cirrhosis.

Background & Aims: Efruxifermin has shown clinical efficacy in patients with non-alcoholic steatohepatitis (NASH) and F1-F3 fibrosis. The primary objective of the BALANCED Cohort C was to assess the safety and tolerability of efruxifermin in patients with compensated NASH cirrhosis. Methods: Patients with NASH and stage 4 fibrosis (n = 30) were randomized 2:1 to receive efruxifermin 50 mg (n = 20) or placebo (n = 10) once-weekly for 16 weeks. The primary endpoint was safety and tolerability of efruxifermin. Secondary and exploratory endpoints included evaluation of non-invasive markers of liver injury and fibrosis, glucose and lipid metabolism, and changes in histology in a subset of patients who consented to end-of-study liver biopsy. Results: Efruxifermin was safe and well-tolerated; most adverse events (AEs) were grade 1 (n = 7, 23.3%) or grade 2 (n = 19, 63.3%). The most frequent AEs were gastrointestinal, including transient, mild to moderate diarrhea, and/or nausea. Significant improvements were noted in key markers of liver injury (alanine aminotransferase) and glucose and lipid metabolism. Sixteen-week treatment with efruxifermin was associated with significant reductions in non-invasive markers of fibrosis including Pro-C3 (least squares mean change from baseline [LSMCFB] -9 µg/L efruxifermin vs. -3.4 µg/L placebo; p = 0.0130) and ELF score (-0.4 efruxifermin vs. +0.4 placebo; p = 0.0036), with a trend towards reduced liver stiffness (LSMCFB -5.7 kPa efruxifermin vs. -1.1 kPa placebo; n.s.). Of 12 efruxifermin-treated patients with liver biopsy after 16 weeks, 4 (33%) achieved fibrosis improvement of at least one stage without worsening of NASH, while an additional 3 (25%) achieved resolution of NASH, compared to 0 of 5 placebo-treated patients. Conclusions: Efruxifermin appeared safe and well-tolerated with encouraging improvements in markers of liver injury, fibrosis, and glucose and lipid metabolism following 16 weeks of treatment, warranting confirmation in larger and longer term studies. Lay summary: Cirrhosis resulting from non-alcoholic steatohepatitis (NASH), the progressive form of non-alcoholic fatty liver disease, represents a major unmet medical need. Currently there are no approved drugs for the treatment of NASH. This proof-of-concept randomized, double-blind clinical trial demonstrated the potential therapeutic benefit of efruxifermin treatment compared to placebo in patients with cirrhosis due to NASH. Clinical Trial Number: NCT03976401.

Myelofibrosis progression grading based on type I and type III collagen and fibrillin 1 expression boosted by whole slide image analysis.

AIMS: The progression of primary myelofibrosis is characterised by ongoing extracellular matrix deposition graded based on 'reticulin' and 'collagen' fibrosis, as revealed by Gomori's silver impregnation. Here we studied the expression of the major extracellular matrix proteins of fibrosis in relation to diagnostic silver grading supported by image analysis. METHODS AND RESULTS: By using automated immunohistochemistry, in this study we demonstrate that the expression of both types I and III collagens and fibrillin 1 by bone marrow stromal cells can reveal the extracellular matrix scaffolding in line with myelofibrosis progression as classified by silver grading. 'Reticulin' fibrosis indicated by type III collagen expression and 'collagen' fibrosis featured by type I collagen expression were parallel, rather than sequential, events. This is line with the proposed role of type III collagen in regulating type I collagen fibrillogenesis. The uniformly strong fibrillin 1 immune signals offered the best inter-rater agreements and the highest statistical correlations with silver grading of the three markers, which was robustly confirmed by automated whole slide digital image analysis using a machine learning-based algorithm. The progressive up-regulation of fibrillin 1 during myelofibrosis may result from a negative feedback loop as fibrillin microfibrils sequester TGF-ß, the major promoter of fibrosis. This can also reduce TGF-ß-induced RANKL levels, which would stimulate osteoclastogenesis and thus can support osteosclerosis in advanced myelofibrosis. CONCLUSIONS: Through the in-situ detection of these extracellular matrix proteins, our results verify the molecular pathobiology of fibrosis during myelofibrosis progression. In particular, fibrillin 1 immunohistochemistry, with or without image analysis, can complement diagnostic silver grading at decent cell morphology.

Stem Cells from Healthy and Tendinopathic Human Tendons: Morphology, Collagen and Cytokines Expression and Their Response to T3 Thyroid Hormone.

The aim of this study was to investigate the effect of triiodothyronine (T3) on tendon specific markers and cytokines expression of stem cells extracted from human tendons. Indeed, thyroid hormones have been reported to be protective factors, maintaining tendons' homeostasis, whereas tendinopathy is believed to be related to a failed healing response. Healthy and tendinopathic human tendons were harvested to isolate tendon stem/progenitor cells (TSPCs). TSPCs obtained from pathological samples showed gene expression and morphological modifications at baseline in comparison with cells harvested from healthy tissues. When cells were maintained in a medium supplemented with T3 (10-6 M), only pathological populations showed a significant upregulation of tenogenic markers (DCN, TNC, COL1A1, COL3A1). Immunostaining revealed that healthy cells constantly released type I collagen, typical of tendon matrix, whereas pathological ones overexpressed and secreted type III collagen, typical of scarred and impaired tissue. Pathological cells also overexpressed pro- and anti-inflammatory cytokines, suggesting an impaired balance in the presence of T3, without STAT3 activation. Moreover, DKK-1 was significantly high in the culture medium of pathological cell cultures and was reversed by T3. This study opens perspectives on the complex biochemical alteration of cells from pathological tendons, which may lead to the chronic disease context with an impaired extracellular matrix.

Degradable microneedle patches loaded with antibacterial gelatin nanoparticles to treat staphylococcal infection-induced chronic wounds.

Infection-induced chronic wounds cause prolonged pains, a high risk of amputation, and even increased mortality in immunocompromised patients. Here we report an antibacterial microneedle (MN) patch, which features high degradability in biological fluids and gelatinase-responsive release of an antibacterial photothermal peptide AMP-Cypate. We first synthesize gelatin nanoparticles (GNPs) and then conjugate the AMP-Cypate to afford composite AMP-Cypate@GNPs. The proteinaceous nanoparticles can responsively release AMP-Cypate in the presence of gelatinase, an enzyme secreted specifically by Staphylococcus aureus (S. aureus). AMP-Cypate@GNPs were then deposited in the tips of MNs fabricated by PVP and recombinant human type III collagen (Col III) to devise the antibacterial MN/AMP-Cypate@GNP patches. When applied to the infection site, MNs break through the epidermis and the stratum corneum, dissolve in the infected dermis, reach the bacterial colony or biofilm, release AMP-Cypate@GNPs, and exert a gelatinase-responsive photothermal therapy under near-infrared (NIR) irradiation to kill the pathogen S. aureus. In a rat model of staphylococcal infection-induced chronic wounds mimicking the condition of diabetic foot ulcer, the antibacterial MN/AMP-Cypate@GNP patches eradiated the bacterial infection and resulted in complete healing of the wounds, proving its potential application in the treatment of chronic wound infections and diabetic foot ulcers.