To examine the activity of matrix metalloproteinases (MMPs) and aggrecanase in control and diseased human articular cartilage, metabolic fragments of aggrecan were detected with monospecific antipeptide antibodies. The distribution and quantity of MMP-generated aggrecan G1 fragments terminating in VDIPEN 341 were compared with the distribution of aggrecanase-generated G1 fragments terminating in NITEGE 373 . Both types of G1 fragments were isolated from osteoarthritic cartilage. The sizes were consistent with a single enzymatic cleavage in the interglobular domain region, with no further proteolytic processing of these fragments. Both neoepitopes were also detected by immunohistochemistry in articular cartilage from patients undergoing joint replacement for osteoarthritis (OA), rheumatoid arthritis (RA), and in cartilage from adults with no known joint disease.In control specimens, the staining intensity for both G1 fragments increased with age, with little staining in cartilage from 22-wk-old fetal samples. There was also an increase with age in the extracted amount of MMP-generated neoepitope in relation to both aggrecan and collagen content, confirming the immunohistochemical results. After the age of 20-30 yr this relationship remained at a steady state. The staining for the MMP-generated epitope was most marked in control cartilage exhibiting histological signs of damage, whereas intense staining for the aggrecanase-generated fragment was often noted in adult cartilage lacking overt histological damage . Intense staining for both neoepitopes appeared in the more severely fibrillated, superficial region of the tissue.Intense immunostaining for both VDIPEN-and NITEGEneoepitopes was also detected in joint cartilage from patients with OA or RA. Cartilage in these specimens was significantly more degraded and high levels of staining for both epitopes was always seen in areas with extensive cartilage damage. The levels of extracted VDIPEN neoepitope relative to collagen or aggrecan in both OA and RA samples were similar to those seen in age-matched control specimens.Immunostaining for both types of aggrecan fragments was seen surrounding the cells but also further removed in the interterritorial matrix. In some regions of the tissue, both neoepitopes were found while in others only one was detected. Thus, generation and/or turnover of these specific catabolic aggrecan fragments is not necessarily coordinated. Our results are consistent with the presence in both normal and arthritic joint cartilage of proteolytic activity against aggrecan based on both classical MMPs and "aggrecanase.

Disruption of the SLN1 gene neither prevents nor reduces the cartilage destruction associated with CIA. Moreover, SLN1 depletion does not prevent the cleavage of the aggrecan Asn341-Phe342 bond.

The predominant form of 5alpha-reductase (5aR) in human scalp is 5aR1. None the less, clinical studies have shown that finasteride, a selective inhibitor of 5aR2, decreases scalp dihydrotestosterone and promotes hair growth in men with androgenetic alopecia. Immunolocalization studies were thus carried out to examine 5aR isozyme distribution within scalp and, in particular, to determine whether 5aR2 might be associated with hair follicles. 5aR2 was localized using both a rabbit polyclonal and a mouse monoclonal antibody. 5aR1 was detected with a mouse monoclonal antibody. The specificity of these reagents was demonstrated both by immunofluorescence and Western blot analyses of COS cells overexpressing human 5aR1 or 5aR2. When cryosections of scalp from men with androgenetic alopecia were stained with antibody against 5aR2, using immunoperoxidase avidin-biotin complex methodology, immunostaining was observed in the inner layer of the outer root sheath and, in more proximal regions of the follicle, in the inner root sheath. Staining was also prominent in the infundibular region of the follicle, with less intense staining extending throughout the granular layer of the epidermis. Some staining was also seen in sebaceous ducts. Similar results were obtained with both the polyclonal and monoclonal 5aR2 antibodies. In contrast, in scalp cryosections stained with antibody to 5aR1, no immunostaining was observed within hair follicles. Intense staining for the type 1 isozyme was, however, detected within sebaceous glands. Our immunolocalization data suggest that the results seen in clinical trials of men with male pattern hair loss treated with finasteride may be due, at least in part, to local inhibition of 5aR2 within the hair follicle.

Background: Dihydrotestosterone mediates androgendependent diseases, such as acne, hirsutism, and androgenetic alopecia. This hormone is produced from testosterone by the 5␣-reductase enzyme. There are 2 isozymes of 5␣-reductase (types 1 and 2) that differ in their localization within the body and even within the skin. Activity of the type 1 isozyme predominates in sebaceous glands, where it may be involved in regulation of sebum production. Since specific inhibition of 5␣-reductase type 1 may represent a novel therapeutic approach to acne, it is important to define the localization of these isozymes in normal sebaceous follicles and acne lesions.Observations: Skin biopsy specimens were obtained from the backs of 11 subjects: 8 with acne and 3 without acne. Sections of normal follicles, open comedones, closed comedones, and inflammatory lesions were incubated with antibodies to types 1 and 2 5␣-reductase. In all samples, the type 1 antibody localized specifically to sebaceous glands, and the type 2 antibody localized to the companion layer of the hair follicle (the innermost layer of the outer root sheath) and granular layer of the epidermis. Localization of the type 2 isozyme was also noted within the walls of open and closed comedones and in endothelial cells from sections of inflammatory lesions. Conclusions:The immunolocalization of 5␣-reductase isozymes in normal sebaceous follicles and acne follicles is similar to the pattern described in terminal hair follicles and corresponds with the findings of biochemical studies that have demonstrated predominance of type 1 activity in sebaceous glands. The function of type 2 5␣reductase in comedones or endothelial cells in inflammatory lesions is unknown.

Given the ongoing challenge of inaccurate survival estimates, this data suggests what may be of greatest clinical utility is to use specific PPS ratings as triggers for key care discussions among patients with incurable and progressive cancer.