Search Results (129)

Type XVII collagen (COL17) is crucial for skin integrity but difficult to produce. To achieve high-level secretory expression, a human COL17 segment was designed and cloned into the pPIC9K vector with six different α-mating factor signal peptides and integrated into Komagataella phaffii GS115. Fed-batch fermentation in a 5 L bioreactor yielded 4.7 g/L of recombinant COL17. Functional assays showed that it promoted the proliferation of human skin fibroblast (HFF) and immortalized keratinocytes (HaCaT), upregulated COL1A1, COL3A1, and TIMP1 in HFF cells, and enhanced skin barrier-related genes (KRT1, KRT5, KRT10, KRT14, IVL, LOR, FLG) in HaCaT cells. In a UVB-induced photoaging model, COL17 reduced reactive oxygen species (ROS) and matrix metalloproteinase 3 (MMP3) activity. This recombinant collagen exhibits photoprotective, regenerative, and barrier-enhancing activities, offering potential for skincare and tissue engineering. Full article

A therapeutic antibody, CSL305, has been developed, which combines inhibition of the complement classical and lectin pathways via complement C2 binding with an ability to act as an antagonist of the neonatal Fc receptor (FcRn). CSL305 binds to human C2 (huC2) but shows no binding or activity against mouse C2 precluding its use in mouse models of disease to fully assess in vivo efficacy. To circumvent this, a mouse strain was developed that replaced the expression of mouse C2 with huC2 by homologous recombination. These mice (huC2 “knock-in”; KI) were shown to express huC2 protein and to have complement activity. Interestingly, male huC2-KI mice showed much stronger complement activity compared to female mice and were also sensitive to inhibition by CSL305. Two models of disease using male huC2-KI mice were then used to assess the in vivo efficacy of CSL305. The first was an attenuated passive anti-glomerular basement membrane (GBM) glomerulonephritis model involving complement activation as its primary mechanism of action. CSL305 showed dose-dependent inhibition of disease as measured by urine albumin, with reductions in kidney cellular infiltration and plasma C3 cleaved fragments C3b/C3c/iC3b also observed. The second model was a collagen autoantibody-induced arthritis (CAIA) mouse model. Here, CSL305 showed a significant and dose-dependent inhibition of clinical score in both prophylactic and therapeutic settings, mediated exclusively via its FcRn mechanism of action. Although the animal models used in this study were found to preclude the demonstration of a synergistic effect on both mechanisms, CSL305 does act in vivo as both a complement inhibitor and as a FcRn antagonist to ameliorate disease. Full article

Periodontitis-induced destruction of periodontal tissues and tooth loss remain major clinical challenges. Although periodontal regenerative therapies aim to reconstruct damaged structures, particularly the repair of alveolar bone defects, current biomaterials have limited capacity to simultaneously promote cell adhesion and osteogenic differentiation. Amelogenin (Am) plays a key role in mineralized tissue development through its highly conserved N-terminal and C-terminal regions, but studies have shown that Am has certain limitations in promoting cell adhesion. In contrast, the arginine-glycine-aspartic acid (RGD) domain of fibronectin (FN) effectively mediates cell–matrix adhesion. Based on these properties, we developed a novel recombinant fusion protein (rtAmR) by combining the conserved regions of Am with the RGD domain of FN. In vitro, rtAmR significantly promoted the adhesion and spreading of human stem cells from the apical papilla (hSCAPs) compared with the control group. Quantitative analysis showed that the number of adherent cells and the cell spreading area in the rtAmR group were 1.9-fold and 2.1-fold higher than those in the rhAm group, respectively. In osteogenic differentiation assays, rtAmR exhibited activity comparable to that of rhAm and even outperformed rhAm in terms of alkaline phosphatase (ALP) activity, collagen type I (COL I) expression, and calcium nodule formation. In a Sprague–Dawley (SD) rat alveolar bone defect model, rtAmR treatment significantly promoted bone regeneration, achieving superior bone volume/total volume (BV/TV) values compared to the rhAm and rhFN groups. Immunohistochemistry revealed that rtAmR did not obviously increase neutrophils, mast cells, or M2 macrophages versus control, confirming its biosafety and suggesting M2-independent osteogenesis. These findings suggest that rtAmR is a promising bifunctional bioactive protein for periodontal bone regeneration. Full article

Critical-sized bone defects represent a major clinical challenge, as defects of this magnitude do not heal spontaneously without regenerative intervention. This study aimed to evaluate the osteoinductive effects of recombinant human bone morphogenetic protein-2 (rhBMP-2) loaded β-tricalcium phosphate (β-TCP) scaffolds on bone regeneration and vascularization in a rabbit calvarial critical-sized defect model. Eighteen male New Zealand White rabbits were used, and four standardized circular defects (5 mm in diameter) were created in the calvaria of each animal. The defects were assigned to four groups: control (unfilled), β-TCP + 5 µg rhBMP-2, β-TCP + 10 µg rhBMP-2, and β-TCP + 20 µg rhBMP-2. Bone healing was evaluated at 2, 4, and 8 weeks using histological, histomorphometric, and cluster of differentiation 31 (CD31) immunohistochemical analyses. The results demonstrated that rhBMP-2–loaded β-TCP scaffolds significantly enhanced bone regeneration compared with the control group, with a progressive increase in bone formation observed with increasing rhBMP-2 doses. The β-TCP + 20 µg rhBMP-2 group exhibited the highest levels of new bone formation, more advanced bone maturation, improved collagen organization, and increased vascularization. However, no statistically significant differences were observed between the 10 µg and 20 µg groups at later time points (p > 0.05), suggesting a dose-dependent saturation (plateau) effect. In conclusion, rhBMP-2–loaded β-TCP scaffolds promote bone regeneration and angiogenesis in a dose-related manner up to a threshold, beyond which additional increases in dose do not result in proportional improvements. These findings emphasize that optimal rhBMP-2 dosing is critical to maximize regenerative outcomes while avoiding unnecessary dose escalation. Full article

Chronic liver diseases, including fibrosis and hepatocellular carcinoma (HCC), are primarily driven by oxidative stress, yet traditional antioxidant therapies often lack the specificity and efficacy required for clinical success. This review evaluates the emerging therapeutic potential of two atypical globins, cytoglobin (CYGB) and neuroglobin (NGB), exploring their unique hexacoordinated heme structures that enable potent reactive oxygen and nitrogen species (ROS/RNS) scavenging and redox-regulated signaling. We summarize a broad range of in vitro and in vivo evidence demonstrating that these globins deactivate hepatic stellate cells, reduce extracellular matrix accumulation, and function as tumor suppressors by modulating pathways such as Raf/MEK/ERK and NRF2. In human cohorts, CYGB expression levels inversely correlate with the progression of Metabolic Dysfunction-Associated Steatohepatitis (MASH) and HCC, highlighting its potential as a clinical biomarker. Furthermore, recombinant protein therapies involving CYGB and NGB show promise in promoting collagen degradation and inhibiting malignant transformation. We conclude that CYGB and NGB represent sophisticated catalytic redox regulators that offer a novel therapeutic paradigm for restoring redox homeostasis. While delivery and pharmacokinetic barriers remain, these globins are highly promising candidates for first-in-class biologics in hepatology. Full article

Heart failure (HF) is the leading cause of morbidity and mortality worldwide, while myocardial fibrosis acts as a pivotal hallmark, which exacerbates ventricular dysfunction and remodeling in HF. In this study, we found FGF23, a critical endocrine regulator, which regulates phosphate and vitamin D metabolism, was significantly upregulated in fibrotic mouse hearts after transverse aortic constriction (TAC). By using the FGF23 monoclonal antibody, we found that inhibition of FGF23 alleviated TAC-induced cardiac fibrosis, while injection of recombinant FGF23 (rFGF23) protein exacerbated tissue fibrosis in mouse hearts after TAC. RNA sequencing indicated that FGF23 may promote cardiac fibroblast proliferation and activation in stressed mouse hearts. In human primary cardiac fibroblasts, rFGF23 treatment further upregulated the expression of Ki67, Cyclin D1, Cyclin E1, PCNA, α-SMA, and collagen 1A1 after TGF-β stimulation. Further results indicated that FGF23 promoted cardiac fibroblast proliferation and activation through FGFR4 and activated the downstream MAPK/ERK signaling. This study suggests a role of FGF23 in the regulation of myocardial fibrosis, which shows the potential of targeting FGF23 in the treatment of HF and cardiac fibrosis. Full article

Collagen is the most abundant structural and functional protein in humans and other vertebrates. It possesses remarkable biological functions and is widely used in food, cosmetics, and healthcare. Currently, mainstream animal-derived collagen materials carry risks such as viral transmission and allergic reactions. However, recombinant collagen, heterologously expressed using genetic recombination technology combined with high-density fermentation processes, offers greater biocompatibility, low immunogenicity, and consistent quality, offering promising development prospects. However, current research on recombinant collagen still faces challenges such as low yield and poor functionality. This article briefly describes the structure, types, and functions of collagen, discusses the advantages and limitations of different recombinant collagen expression systems, and highlights the strategies for improving the yield and optimizing the function of recombinant collagen, ranging from gene editing to fermentation optimization. In highlighting practical approaches to achieving high yield, we present a series of case examples to illustrate the successful application of these principles. This review aims to help researchers, engineers, and industry practitioners better understand research trends in the expression and production of recombinant collagen, and to promote its further development and commercialization across diverse application areas. Full article

Collagen is the primary protein in the extracellular matrix of human cells and the body and is essential for cell structure and function. Here, for the first time, we report a method for producing recombinant triple-helical collagen type III (rhCOL3) in transgenic tobacco as a bioreactor. We constructed a pMDV-COL3A1 vector containing the human type III collagen gene COL3A1, as well as a pMDV-COL3A1:5E vector that coexpressed COL3A1 and the enzymes required for its posttranslational modification. These two vectors were used to transform tobacco genetically. The COL3A1 gene was successfully coexpressed in tobacco plants with four enzymes that promote its posttranslational modification. The transcriptional level of COL3A1 in the transgenic lines coexpressing posttranslational modification genes was greater than that in the transgenic lines expressing only COL3A1. The enzyme-modified recombinant collagen was subsequently purified from a COL3A1:5E transgenic line. Our experimental results demonstrated that the terminal propeptides of plant-derived rhCOL3 can be correctly cleaved through the enzymatic hydrolysis of procollagen by coexpressed procollagen C proteinase (PCP) and procollagen N proteinase (PNP). The plant-derived rhCOL3 was thermally stable because the purified peptide chains can form a triple helix structure. Experiments have shown that plant-derived rhCOL3 has biological activity. In this study, functional recombinant full-length mature type III collagen with a triple-helix structure was successfully expressed in tobacco, providing a foundational plant-made material for future applications of collagen in human skin and bone repair in regenerative medicine. Full article

Background: Collagen XIα1, encoded by the COL11A1 gene, is a minor fibrillar collagen that is overexpressed in various human cancers, in which its presence correlates with tumor aggressiveness and progression. Methods: In this study, we developed two novel mouse monoclonal antibodies (mAbs)—anti-colXIα1 clone 3 and anti-colXIα1 clone 9—that target the putative C-telopeptide of human collagen XIα1. These antibodies target the RRHTEGMQA sequence, a unique nine-amino-acid stretch within the putative C-telopeptide of human collagen XIα1. Results: Corresponding to nearly identical V(D)J gene segments and complementarity-determining regions (CDRs), the antibodies specifically bound the RRHTEGMQA epitope in ELISAs but did not react with the C-propeptide. This specificity was further confirmed with the purified anti-colXIα1 clone 9 mAb, which demonstrated strong reactivity against recombinant proteins containing the RRHTEGMQA sequence in both ELISAs and Western blot assays. This sequence seems to behave as a linear B-cell neoepitope, in which the RRHT motif is crucial for epitope recognition. Otherwise, no immunodetections were observed, either in cultures and lysates from the COL11A1-highly expressing A204 human cell line or on tissue sections from specimens of human pancreatic ductal adenocarcinoma (PDAC), with strong desmoplastic reactions. Conclusions: Given the lack of precise knowledge of the characteristics of the putative C-telopeptide of human collagen XIα1, the presented antibodies could enhance our understanding of the processing of human procollagen XIα1 and contribute to better characterization of the tumor microenvironment of COL11A1-expressing cancers. Full article

Background: Delayed or non-union fractures comprise 5–10% of cases, indicating the need for biologic interventions. Recombinant human bone morphogenetic protein-2 (rhBMP-2) is a potent osteoinductive agent; yet, collagen carrier-based uncontrolled release causes adverse events. We evaluated the safety and efficacy of a hydroxyapatite (HA) carrier-based rhBMP-2 delivery system for acute traumatic upper and lower fractures exhibiting bone defects. Methods: This prospective, multicenter, single-arm clinical trial enrolled 90 patients who underwent surgery using a hydroxyapatite (HA) carrier-based rhBMP-2 delivery system (Novosis^TM). Radiographically validated union at 6 and 12 months post-surgery and treatment success (union without additional surgery) were used to assess efficacy. The incidence, type, and severity of all device-related adverse events during follow-up were monitored by investigators to evaluate safety. Results: Of the 90 patients enrolled, 81 were included in the full analysis set. The mean age was 58.5 years, and 18.6% (15/81) had open fractures. At 6 months post-surgery, radiographically validated union was achieved in 81.5% (66/81) of patients, increasing to 96.2% (77/81) at 12 months after surgery. Treatment success was 95.0% (76/81). Adverse events were rare (1/81, 1.2%). No ectopic ossification, systemic complications, or severe inflammatory responses were observed. Conclusions: HA-based rhBMP-2 intervention demonstrated favorable union rates and safety with minimal complications in acute upper and lower fractures with bone defects. The biocompatibility and controlled-release properties of HA likely improved efficacy and reduced complications. Results should be interpreted as feasibility data from a heterogeneous case series without a control group. Larger randomized controlled comparative trials are warranted for optimal dosing and evaluating efficacy and cost-effectiveness. Full article

Collagen types I and III are primary structural proteins that maintain human skin integrity, and their ratio is disrupted during aging. In this study, we developed a biomimetic 3D skin model with vascularization potential to evaluate the effects of nano-zinc oxide (nano-ZnO) in a physiologically relevant context. The model used methacrylated recombinant human collagen (RHC-MA) bioinks with tunable collagen I/III ratios that mimic the skin of children and adults. The bioinks exhibited excellent printability and mechanical properties that enabled the 3D bioprinting of full-thickness skin products, including dermal layers with human skin fibroblasts (HSFs) and human umbilical vein endothelial cells (HUVECs), as well as epidermal layers with keratinocytes (KCs). The model recapitulated native skin architecture, and key markers such as keratin 10 (K10), keratin 14 (K14), and cluster of differentiation 31 (CD31) were determined. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis showed that nano-ZnO significantly modulated genes associated with apoptosis, inflammation, and oxidative stress in skin cells. Full article

FAM20C, previously known as Golgi casein kinase (GCK), is a serine/threonine kinase localized to the Golgi apparatus and classified within the acidophilic kinase family. Its phosphorylation motif is characterized by a glutamic acid residue at the +2 position relative to the target site. Before its molecular identity was established, analysis of a limited number of phosphosites in secreted proteins showed that around 70% matched the GCK consensus sequence, suggesting that GCK is the principal kinase for secreted proteins. Following the identification of GCK as FAM20C, the generation of FAM20C knockout cell lines and phosphoproteomic data confirmed its role: approximately 80% of serine/threonine phosphosites in the secretome of two different human cell lines were shown to depend on FAM20C. In this study, comparative analysis of in vitro phosphorylation datasets from a broad panel of recombinant Ser/Thr kinases confirmed that the FAM20C consensus sequence is distinct from those of other acidophilic kinases. Examination of experimentally identified human phosphosites within the secretory pathway revealed strong conservation of the FAM20C consensus, firmly establishing this enzyme as the master Ser kinase of the entire pathway. From this dataset, we defined the putative FAM20C substratome, comprising 443 phosphosites across 256 proteins, ~77% of which had not been previously linked to FAM20C. This represents the most extensive FAM20C substratome to date and a valuable resource for functional studies. Notably, enrichment analysis highlights strong links between FAM20C and major extracellular pathways, including collagen fibril organization, complement activation, and blood coagulation, underscoring an underappreciated role for this kinase in regulating hemostasis and innate immunity. Full article

Reconstruction of large and complex hard tissue defects remains a major clinical challenge, as conventional autografts and allografts are often limited in availability, biological compatibility, and long-term efficacy, particularly for extensive defects or poor bone quality. Recombinant human bone morphogenetic protein-2 (rhBMP-2) is a potent osteoinductive factor capable of initiating the complete cascade of bone formation. However, its clinical use is restricted by dose-dependent complications such as inflammation, ectopic ossification, and osteolysis. This review synthesizes current evidence on the safety profile of rhBMP-2 and examines strategies to enhance its therapeutic index. Preclinical and clinical data indicate that conventional collagen-based carriers frequently cause rapid burst release and uncontrolled diffusion, aggravating adverse outcomes. It is noteworthy that low doses of rhBMP-2 (0.5–0.7 mg/level in anterior cervical discectomy and fusion (ACDF) or 0.5–1.0 mg/level in transforaminal lumbar interbody fusion (TLIF)) provide the optimal balance of efficacy and safety. Advanced biomaterial-based platforms, such as bioceramic–polymer composites, injectable hydrogels, and 3D-printed scaffolds, enable spatially and temporally controlled release while maintaining osteogenic efficacy. Molecular delivery approaches, including chemically modified messenger RNA (cmRNA) and regional gene therapy, provide transient, site-specific rhBMP-2 expression with reduced dosing and minimal systemic exposure. By integrating mechanistic insights with translational advances, this review outlines a framework for optimizing rhBMP-2-based regenerative protocols, emphasizing their potential role in multidisciplinary strategies for reconstructing complex hard tissue defects. Full article

Recombinant human bone morphogenetic protein-2 (rhBMP-2) is widely used to promote bone regeneration. However, conventional surface-attached delivery on absorbable collagen sponges causes a rapid burst release, excessive inflammation, and suboptimal healing. To overcome these limitations, we developed a thermally controlled Poly(DL-lactide-co-glycolide) (P_DLLGA) encapsulation system, designed to stabilize rhBMP-2 and enable controlled release. rhBMP-2 was incorporated in P_DLLGA pellets using the hot-melt extrusion of a lyophilized mixture containing poloxamer 407 and hydroxypropyl-β-cyclodextrin, and terminal sterilization (X-ray irradiation). The released rhBMP-2 maintained its molecular integrity after sterilization and remained stable for up to 732 days in storage, as confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and capillary electrophoresis (CE). Further, high-affinity binding between released rhBMP-2 and BMPR-IA was confirmed by bio-layer interferometry (BLI), and the released protein induced a robust in vitro ALP response, confirming preserved osteogenic activity. Our encapsulation approach for rhBMP-2 using P_DLLGA, including the combination product with β-TCP (LDGraft; Locate Bio, Nottingham, UK), provides a stable and bioactive rhBMP-2 delivery strategy with inherent dose-shielding properties, supporting safe, controlled, and effective bone regeneration therapies. Full article

Type II collagen (Col2), a crucial structural protein in hyaline cartilage, is essential for cartilage integrity and facilitating injury repair. However, research on recombinant type II collagen still faces many challenges, such as structure and yield, which limit the application of recombinant Col2 in biomedical fields. In this study, we achieved high-yield expression of full-length human Col2 (rhCol2) in CHO cells. The physical and chemical properties of rhCol2 were very close to native Col2, including molecular weight, triple helix structure, thermal stability and self-assembly capacity. Functional assays of primary chondrocytes have demonstrated that rhCol2 can effectively promote chondrocyte proliferation and increase the expression levels of cartilage-specific genes (Col2a1, Aggrecan, and Sox-9). Moreover, a cartilage defect model was surgically created in SD rats demonstrated that rhCol2 significantly enhanced cartilage repair, and the severity of the defect was assessed through histological and micro-CT analyses. Human chondrocytes were utilized to compare the effects of different collagens and verified through a series of functional experiments. In conclusion, these findings indicate that rhCol2 is an effective biomaterial and is expected to promote the application of recombinant collagen in the field of cartilage repair. Full article