Production and Characterization of Recombinant Rat Non-Collagen Domain of α3 Chain of Type IV Collagen α3 (IV) NC1 Antigen

The glomerulonephritis disease is characterized by inflammation of glomeruli or small blood vessels in the kidney that causes kidney diseases. The reason of glomerulonephritis disease is to deposit the anti-GBM auto antibody in the glomerular basement membrane. The type IV collagen is the main component of glomerular basement membrane that has α3 chain of type (IV) collagen of non-collagenous domain which contains N-terminal 7S domain, a triple helical collagenous domain and C-terminal non-collagenous glomerular domain (NC1). The amino terminal of α3 (IV) NC1 that induces the Experimental Autoimmuno Glomerulonephritis (EAG) in rat model has been identified. The recombinant rat α3 (IV) NC1 antigen has nine amino acid spans that are consistent with antibody or T cell epitope that induces in EAG. The research is carried out on the recombinant rat α3 (IV) NC1 production, purification, quantification, and characterization. The circulation of anti-GBM antibody in glomerular basement membrane can be measured by the ELISA assay. In addition, the recombinant rat antigen is secreted in HEK293 cell supernatant that is purified by Anti-FLAG M2 monoclonal IgG antibody affinity column and characterized and quantified by SDS-PAGE gel electrophoresis and Western blotting techniques.

NC1-peptide derived from collagenα3(IV)chain is a blood-tissue barrier regulator:lesson from the testis

Collagenα3(IV)chains are one of the major constituent components of the basement membrane in the mammalian testis.Studies have shown that biologically active fragments,such as noncollagenase domain(NC1)-peptide,can be released from the C-terminal region of collagenα3(IV)chains,possibly through the proteolytic action of metalloproteinase 9(MMP9).NC1-peptide was shown to promote blood-testis barrier(BTB)remodeling and fully developed spermatid(e.g.,sperm)release from the seminiferous epithelium because this bioactive peptide was capable of perturbing the organization of both actin-and microtubule(MT)-based cytoskeletons at the Sertoli cell-cell and also Sertoli-spermatid interface,the ultrastructure known as the basal ectoplasmic specialization(ES)and apical ES,respectively.More importantly,recent studies have shown that this NC1-peptide-induced effects on cytoskeletal organization in the testis are mediated through an activation of mammalian target of rapamycin complex 1/ribosomal protein S6/transforming retrovirus Akt1/2 protein(mTORC1/rpS6/Akt1/2)signaling cascade,involving an activation of cell division control protein 42 homolog(Cdc42)GTPase,but not Ras homolog family member A GTPase(RhoA),and the participation of end-binding protein 1(EB1),a microtubule plus(+)end tracking protein(+TIP),downstream.Herein,we critically evaluate these findings,providing a critical discussion by which the basement membrane modulates spermatogenesis through one of its locally generated regulatory peptides in the testis.

Type XIX collagen: a promising biomarker from the basement membranes

Among collagen members in the collagen superfamily,type XIX collagen has raised increasing interest in relation to its structural and biological roles.Type XIX collagen is a Fibril-Associated Collagen with Interrupted Triple helices member,one main subclass of collagens in this superfamily.This collagen contains a triple helix composed of three polypeptide segments aligned in parallel and it is associated with the basement membrane zone in different tissues.The molecular structure of type XIX collagen consists of five collagenous domains,COL1 to COL5,interrupted by six non-collagenous domains,NCI to NC6.The most relevant domain by which this collagen exerts its biological roles is NCI domain that can be cleavage enzymatically to release matricryptins,exerting anti-tumor and anti-angiogenic effect in murine and human models of cancer.Under physiological conditions,type XIX collagen expression decreases after birth in different tissues although it is necessary to keep its basal levels,mainly in skeletal muscle and hippocampal and telencephalic interneurons in brain.Notwithstanding,in amyotrophic lateral sclerosis,altered transcript expression levels show a novel biological effect of this collagen beyond its structural role in basement membranes and its anti-tumor and anti-angiogenic properties.Type XIX collagen can exert a compensatory effect to ameliorate the disease progression under neurodegenerative conditions specific to amyotrophic lateral sclerosis in transgenic SOD1 G93 A mice and amyotrophic lateral sclerosis patients.This novel biological role highlights its nature as prognostic biomarker of disease progression in and as promising therapeutic target,paving the way to a more precise prognosis of amyotrophic lateral sclerosis.