Procollagen Synthesis Research Articles

Natural Arctium lappa fruit extract improves the clinical signs of aging skin

Subclinical, chronic tissue inflammation involving the generation of cytokines (e.g., interleukin-6 and tumor necrosis factor-alpha) might contribute to the cutaneous aging process. This study aims to screen for an active ingredient with anti-inflammatory (i.e., reduction of interleukin-6 and tumor necrosis factor-alpha) and matrix-stimulating efficacy which improves the clinical signs of skin aging in vivo. In vitro studies with pure Arctiin were performed investigating the inhibition of cytokine induction and stimulation of collagen neo-synthesis. In vivo home-in-use studies using an Arctium lappa fruit extract-containing formulation were carried out to determine procollagen and hyaluronan synthesis, hyaluronan synthase-2 gene expression, and reduction of wrinkle volume after treatment. In vitro studies on human dermal fibroblasts and monocyte-derived dendritic cells supplemented with pure Arctiin showed relative to untreated control cells a stimulation of collagen synthesis and a decrease in interleukin-6 and tumor necrosis factor-alpha concentration, respectively. In addition, topical in vivo application of an A. lappa fruit extract-containing formulation for 12 weeks significantly stimulated procollagen synthesis and increased hyaluronan synthase-2 expression as well as hyaluronan levels compared to vehicle-treated control areas. Similarly, after a 4-week treatment with an A. lappa fruit extract-containing formulation, wrinkle volume in the crow's feet area was significantly reduced as compared to treatment with the vehicle. Our data show that topical treatment with a natural A. lappa fruit extract significantly improves the metabolism of the dermal extracellular matrix and leads to a visible wrinkle reduction in vivo. In conclusion, A. lappa fruit extract represents a targeted means to regenerate dermal structures and, thus, offers an effective treatment option for mature skin.

Mechanisms of carvacrol-induced expression of type I collagen gene

Skin aging is accompanied by wrinkle formation and appears to be principally related to decreases in the levels of type I collagen, the primary component of the dermal layer of skin. To investigate the effect of carvacrol on collagen gene expression and its mechanisms of action. To elucidate the effect of carvacrol on collagen expression and its mechanism, several experiments were performed in human dermal fibroblasts. Collagen production, small interference RNA, Ca(2+) mobilization, COL1A2/AP-1 luciferase reporter assays and Western blots for proteins that are involved in collagen gene expression were used in this study. Carvacrol activated both the human COL1A2 promoter activity and the synthesis of human type I procollagen. Additionally, we attempted to characterize the mechanism of action of carvacrol in type I procollagen synthesis. In a human COL1A2 promoter luciferase assay, the small interference RNA for SP-1 did not reduce the carvacrol-induced promoter activation. Also, Smad 2 phosphorylation was not induced by carvacrol. However, in the AP-1 (activator protein-1) luciferase reporter assay and in the Western blot analysis for mitogen-activated protein kinases (MAPKs), carvacrol induced the activation of the AP-1 promoter and the phosphorylation of JNK and ERK1/2 (p42/44 MAPK), but did not induce the phosphorylation of p38 MAPK. Carvacrol also induced intracellular Ca(2+) mobilization and phosphorylation of phospholipase Cgamma1 (PLCgamma1), a molecule upstream of Ca(2+). In addition, PAO, a PLCgamma1 inhibitor, attenuated the carvacrol-induced production of collagen. Our study suggests that the PLCgamma1 signaling pathway may be involved in the carvacrol-induced expression of the type I collagen gene.

Intracellular Localization of Cthrc1 Characterizes Differentiated Smooth Muscle

We recently reported expression of collagen triple helix repeat containing-1 (Cthrc1) in injured arteries and proteolytic cleavage of Cthrc1 in smooth muscle cells in vitro. The present study characterizes Cthrc1 processing and determines its biological significance. Domain-specific antibodies localized full-length Cthrc1 in the cytoplasm of vascular, gastrointestinal, and uterine smooth muscle as well as in some neurons. Unlike smooth muscle alpha-actin, Cthrc1 was not expressed in the embryonic myocardium. Intracellular localization of full-length Cthrc1 was sharply reduced in dedifferentiated smooth muscle of the developing neointima despite the previously shown increase in mRNA levels with accompanying extracellular Cthrc1 immunoreactivity. Immunoblotting suggested an apparent covalent association of monomeric full-length Cthrc1 with a cytoplasmic protein present in differentiated smooth muscle. Plasmin was identified as a protease that cleaved a putative propeptide generating an N-terminally truncated form of Cthrc1 with increased inhibitory activity of procollagen synthesis. Our data show that the differentiated smooth muscle cell phenotype is associated with the intracellular localization of noncleaved Cthrc1 despite the presence of a signal peptide. On arterial injury, increased Cthrc1 expression with apparent extracellular localization of N-terminally truncated Cthrc1 occurs. Removal of the propeptide correlated with increased activity of the molecule.

Periodontal Ehlers–Danlos syndrome associated with type III and I collagen deficiencies

In 1997, Ehlers-Danlos syndrome type VIII (EDS-VIII) was excluded from the diagnostic categories as there were insufficient data regarding the clinical features and the precise underlying molecular basis. However, a recent review of published cases shows that EDS-VIII has distinctive clinical features, which would suggest that it should be considered as a separate entity in future nosological classifications. The underlying molecular defect in EDS-VIII is unknown. A reduction of collagen type III was reported in a single case, but no consistent biochemical or structural changes are detectable. We report a patient with EDS-VIII who showed a reduction in the collagen type I and type III synthesis rates. Type I and type III procollagen and collagen synthesis and secretion rates were investigated in cultured fibroblasts and compared with five healthy controls and three patients with EDS type IV whose fibroblasts were cultured in parallel.

MDI 301, a nonirritating retinoid, improves abrasion wound healing in damaged/atrophic skin

MDI 301 is a picolinic acid-substituted ester of 9-cis retinoic acid. It has been shown in the past that MDI 301 increases epidermal thickness, decreases matrix metalloproteinase (MMP) activity, and increases procollagen synthesis in organ-cultured human skin. Unlike all-trans retinoic acid (RA), MDI 301 does not induce expression of proinflammatory cytokines or induce expression of leukocyte adhesion molecules in human skin. In the present study we examined topical MDI 301 treatment for ability to improve the structure and function of skin in three models of skin damage in rodents and for ability to improve abrasion wound healing in these models. MDI 301 was applied daily to the skin of rats treated with the potent corticosteroid, clobetasol propionate, to the skin of diabetic rats (8 weeks posttreatment with streptozotocin) and to the skin of aged (14-16-month-old) rats. In all three models, subsequently induced abrasion wounds healed more rapidly in the retinoid-treated animals than in vehicle-treated controls. Immediately after complete wound closure, tissue from the wound site (as well as from a control site) was put into organ culture and maintained for 3 days. At the end of the incubation period, culture fluids were assessed for soluble type I collagen and for MMPs-2 and -9. In all three models, the level of type I collagen was increased and MMP levels were decreased by MDI 301. In all three models, skin irritation during the retinoid-treatment phase was virtually nonexistent.

Fermentable metabolite of Zymomonas mobilis controls collagen reduction in photoaging skin by improving TGF-β/Smad signaling suppression

Solar ultraviolet (UV) irradiation causes damages on human skin and premature skin aging (photoaging). UV-induced reduction of type I collagen in dermis is widely considered primarily induction of wrinkled appearance of photoaging skin. Type I procollagen synthesis is reduced under UV irradiation by blocking transforming growth factor-beta (TGF-beta)/Smad signaling; more specifically, it is down-regulation of TGF-beta type II receptor (T beta RII). Therefore, preventing UV-induced loss of T beta RII results decreased type I collagen reduction in photoaging skin. Zymomonas mobilis is an alcohol fermentable, gram-negative facultative anaerobic bacterium whose effect on skin tissue is scarcely studied. We investigated the protective effects of fermentable metabolite of Z. mobilis (FM of Z. mobilis) against reduction of type I procollagen synthesis of UV-induced down-regulation of T beta RII in human dermal fibroblasts FM of Z. mobilis was obtained from lyophilization of bacterium culture supernatant. The levels of T beta RII and type I procollagen mRNA in human dermal fibroblasts were measured by quantitative real-time RT-PCR, and T beta RII protein levels were assayed by western blotting. T beta RII, type I procollagen, and type I collagen proteins in human dermal fibroblasts or hairless mouse skin were detected by immunostaining. FM of Z. mobilis inhibited down regulation of T beta RII mRNA, and protein levels in UVB irradiated human dermal fibroblasts consequently recover reduced type I procollagen synthesis. These results indicate UVB irradiation inhibits type I procollagen synthesis by suppression of TGF-beta/Smad signaling pathway, and FM of Z. mobilis has inhibitory effect on UVB-induced reduction of type I procollagen synthesis. While short period UVB irradiation decreased both T beta RII and type I procollagen protein levels in hairless mouse skin, topical application of FM of Z. mobilis prevented this decrease. Wrinkle formation in hairless mouse skin surface was accelerated by continuous 5 month UVB irradiation along with a reduction of type I collagen in the dermis, but this change was prevented by topical application of FM of Z. mobilis. From this experimental data, it is suggested that FM of Z. mobilis is effective for suppression of wrinkle formation in photoaging skin by inhibition of type I procollagen synthesis reduction.

Inhibition effects of constituents of Radix Salviae Miltiorrhizae on proliferation and procollagen transcription of dermal fibroblasts in systemic sclerosis

To study the effects of tanshinone IIA, danshensu, lithospermate, and protocatechualdehyde on proliferation and procollagen mRNA synthesis of dermal fibroblasts from patients with systemic sclerosis (SSc). Dermal fibroblasts were obtained from 5 patients with SSc, 2 males and 3 females, and sex and age-matched 5 patients undergoing plastic surgery, cultured, and then co-cultured with tanshinone IIA, danshensu, lithospermate, and protocatechualdehyde respectively. MTT test was used to study the effects of the four constituents on fibroblasts cell proliferation. RT-PCR was used to detect the level of procollagen mRNA expression in the control fibroblasts and in the SSc fibroblasts before and after the addition of the four constituents. The 4 constituents all suppressed the proliferation of fibroblasts from SSc patients, with tanshinone IIA being the most effective. The 4 constituents remarkably inhibited the mRNA expression of type I and III procollagen of the fibroblasts with tanshinone II A showing the most inhibition effect that reduced the expressions of procollagen I mRNA by 47% and procollagen III mRNA by 45%. Of the four constituents, tanshinone IIA is the most suppressive effect on the cell proliferation and type I and type III procollagen synthesis of SSc fibroblasts, demonstrating that it has a more antifibrotic effect.

Anti‐wrinkling effects of the mixture of vitamin C, vitamin E, pycnogenol and evening primrose oil, and molecular mechanisms on hairless mouse skin caused by chronic ultraviolet B irradiation

Naturally occurring antioxidants were used to regulate the skin damage caused by ultraviolet (UV) radiation because several antioxidants have demonstrated that they can inhibit wrinkle formation through prevention of matrix metalloproteinases (MMPs) and/or increase of collagen synthesis. We examined the effect of oral administration of the antioxidant mixture of vitamin C, vitamin E, pycnogenol, and evening primrose oil on UVB-induced wrinkle formation. In addition, we investigated the possible molecular mechanism of photoprotection against UVB through inhibition of collagen-degrading MMP activity or through enhancement of procollagen synthesis in mouse dorsal skin. Female SKH-1 hairless mice were orally administrated the antioxidant mixture (test group) or vehicle (control group) for 10 weeks with UVB irradiation three times a week. The intensity of irradiation was gradually increased from 30 to 180 mJ/cm2. Microtopographic and histological assessment of the dorsal skins was carried out at the end of 10 weeks to evaluate wrinkle formation. Western blot analysis and EMSA were also carried out to investigate the changes in the balance of collagen synthesis and collagen degradation. Our antioxidant mixture significantly reduced UVB-induced wrinkle formation, accompanied by significant reduction of epidermal thickness, and UVB-induced hyperplasia, acanthosis, and hyperkeratosis. This antioxidant mixture significantly prevented the UVB-induced expressions of MMPs, mitogen-activated protein (MAP) kinase, and activation of activator protein (AP)-1 transcriptional factor in addition to enhanced type I procollagen and transforming growth factor-beta2 (TGF-beta2) expression. Oral administration of the antioxidant mixture significantly inhibited wrinkle formation caused by chronic UVB irradiation through significant inhibition of UVB-induced MMP activity accompanied by enhancement of collagen synthesis.

TGF‐β–induced fibrosis and SMAD signaling: oligo decoys as natural therapeutics for inhibition of tissue fibrosis and scarring

Transforming-growth factor-beta (TGF-beta) is a pleiotrophic growth factor that is synthesized by many cells in the body. This growth factor is chemotactic for fibroblasts, stimulates fibroblast proliferation, and increases the synthesis of a number of extracellular matrix proteins including collagens. The TGF-beta activator protein is a transacting factor, which binds to the TGF-beta element in the distal promoter of the COL1A1 collagen gene and induces transcription of this gene. Although transient TGF-beta 1 activity participates in repair and regeneration of tissues, persistent TGF-beta 1 function affects excessive fibrosis and ultimately scarring of both skin and internal organs. Scarring of internal organ (e.g., liver and lung) results in a loss of function and ultimately death may occur. The central issue of this review is that phosphorothioate double-stranded decoys or other decoys decrease procollagen gene expression, procollagen synthesis, and collagen during fibrogenesis. The rationale is that the decoys containing the TGF-beta element or other gene transcription regulatory CIS-elements bind the transacting proteins preventing the latter from binding to the CIS-element in the 5'-flanking region of the natural gene resulting in transcription inhibition. We will, in part, focus on aspects involved in TGF-beta 1-induced fibrosis that occur during fibrogenesis and the use of the dsTGF-beta element containing oligodeoxynucleotide decoys to control excessive collagen synthesis, and deposition resulting from persistent TGF-beta. In our model of regulation of collagen synthesis, these double-stranded oligo decoys act as promoter competitors, binding to the activator protein either in the cytoplasm or in the nucleus. The significance of the proposed studies is that these novel natural antifibrotics will mimic the effect of glucocorticoids on collagen synthesis during fibrogenesis without the unwanted side effects of these steroids. Based on our previous studies on the molecular mechanisms by which glucocorticoids selectively decrease collagen synthesis, designed phosphorothioate oligodeoxynucleotides resistant to nuclease action will mimic the effects of glucocorticoids at the molecular, cellular, and in vivo levels of collagen synthesis. However, the glucocorticoids significantly inhibit noncollagen protein synthesis. Both the single-stranded and double-stranded oligodeoxynucleotide specifically decrease collagen synthesis without an inhibitory effect on noncollagen protein synthesis. In this review, we will specifically ask if TGF-beta-induced collagen synthesis is inhibited in cell culture and in vivo by using the double-stranded oligodeoxynucleotide decoys, will this inhibit fibrogenesis and ultimately scarring?

Effect of Camellia japonica oil on human type I procollagen production and skin barrier function

Type I collagen is the primary component of the skin dermis. Both the quantity and quality of extracellular collagen are primarily related to skin ageing. In this study, we investigated the possibility that Camellia japonica oil (CJ oil) may be introduced as an anit-wrinkle agent. As a first step to this end, human COL1A2 promoter luciferase assay was performed in human dermal fibroblast cells. CJ oil was determined to activate human COL1A2 promoter in a concentration-dependent manner. In consistency with this result, while matrix metalloproteinase (MMP)-1 activity was inhibited by CJ oil, human type I procollagen synthesis was also induced by CJ oil. These results suggest the possibility that CJ oil may be involved in the skin ageing. For the evaluation of CJ oil's safety and efficiency on human skin, human skin primary irritation test and trans-epidermal water loss (TEWL) were performed. Transepidermal water loss (TEWL) was measured before treatment then, 1 h and 2 h after treatment; the forearm site was selected to measure TEWL. Also, a human skin primary irritation test was performed on the normal skin (upper back) in 30 volunteers to see if a certain material included in CJ oil has irritation or sensitization potential. In these assays, CJ oil reduced trans-epidermal water loss (TEWL) and did not induce any adverse reactions. Therefore, based on these results, we suggest the possibility that CJ oil may be considered as possible wrinkle-reducing candidates for topical application.

Genetic and Constitutional Influences on Bone Turnover Markers: A Study of Male Twin Pairs

Biochemical markers of bone turnover originating from type I procollagen synthesis or type I collagen breakdown were examined in men using a classic twin study design based on monozygotic (MZ) and dizygotic (DZ) twins. The aim was to estimate the influence of heredity (genes and shared family childhood elements) and constitutional factors in determining procollagen type I amino-terminal propeptide (PINP), type I collagen carboxy-terminal telopeptide (ICTP), and urinary amino-terminal type I collagen telopeptide (NTx) marker levels in a sample of in 98 MZ and 108 DZ male twin pairs. We are not aware of any prior studies conducted in men that address the influence of genetic factors on bone turnover marker variability. The findings support a dominant role for heredity in the variation of bone resorption marker levels in men, with additive genetic effects explaining two-thirds of the variance in the bone resorption markers NTx and ICTP. Genetic factors may contribute less for PINP, a marker of bone formation. The genetic loci influencing PINP or NTx and body weight/disc axial area, although related in part, appeared to be largely independent, indicating that genetic effects on bone turnover are unlikely to be to a large degree a result of genetic regulation of individual body weight.

Stimulation of collagen biosynthesis by topically applied vitamin C

In the dermis, vitamin C is required for the formation of hydroxyprolyl residues to form stable triple-helical collagen molecules, and hydroxylysyl residues operating in crosslinks synthesis. In fibroblasts culture, vitamin C is also known to stabilize the collagen messenger RNAs (m-RNA) and increase procollagen synthesis. The activity in the dermis of vitamin C topically applied on normal human skin is demonstrated in the present study by observing an increased steady-state level of the m-RNA for the two major collagen molecules (I and III) in skin, three of their postradiational processing enzymes (Nand C-procollagen peptidases and lysyloxidase) and markers of dermal and epidermal cells activity.

Procollagen Synthesis is Increased in Hypothyroid Rat Ovary by a Parallel and Compensatory Pathway

Collagen biosynthesis is a multistep process that starts with the transcription and translation of the individual collagen gene. It is characterized by the presence of a large number of co- and posttranslational modifications. Hydroxylysine is found only in animal proteins and mostly in collagens. Procollagen lysyl hydroxylation is the first step in collagen biosynthetic pathway and lysyl hydroxylases (Plod isoforms) are responsible for this enzymatic process. Previously we showed the down regulation of Plod isoforms in hypothyroid ovary. As hypothyroidism is a stress for normal animals, we wanted to explore whether any compensatory pathway exists to balance the reduced lysyl hydroxylation of collagen in hypothyroid rat ovary. In this report we have shown that procollagen I and III are increased in hypothyroid condition and subsequently decreased upon T₃ add-back. Heat Shock Protein-47 is a collagen-specific molecular chaperone and its existence in ovary has been documented. The genes encoding HSP-47, prolyl-4-hydroxylase-α and -β (P4H-?α and -?β) are increased in hypothyroid condition. Down regulation of lysyl hydroxylase in hypothyroid condition results less collagen formation. At the same time over production of procollagens, HSP-47 and P4H is very significant as they may compensate the damage whatsoever caused due to hypothyroidism in ovarian tissue.

Procollagen Synthesis is Increased in Hypothyroid Rat Ovary by a Parallel and Compensatory Pathway

Collagen biosynthesis is a multistep process that starts with the transcription and translation of the individual collagen gene. It is characterized by the presence of a large number of co- and posttranslational modifications. Hydroxylysine is found only in animal proteins and mostly in collagens. Procollagen lysyl hydroxylation is the first step in collagen biosynthetic pathway and lysyl hydroxylases (Plod isoforms) are responsible for this enzymatic process. Previously we showed the down regulation of Plod isoforms in hypothyroid ovary. As hypothyroidism is a stress for normal animals, we wanted to explore whether any compensatory pathway exists to balance the reduced lysyl hydroxylation of collagen in hypothyroid rat ovary. In this report we have shown that procollagen I and III are increased in hypothyroid condition and subsequently decreased upon T₃ add-back. Heat Shock Protein-47 is a collagen-specific molecular chaperone and its existence in ovary has been documented. The genes encoding HSP-47, prolyl-4-hydroxylase-α and -β (P4H-?α and -?β) are increased in hypothyroid condition. Down regulation of lysyl hydroxylase in hypothyroid condition results less collagen formation. At the same time over production of procollagens, HSP-47 and P4H is very significant as they may compensate the damage whatsoever caused due to hypothyroidism in ovarian tissue.

Alpha-Ketoglutarate Stimulates Procollagen Production in Cultured Human Dermal Fibroblasts, and Decreases UVB-Induced Wrinkle Formation Following Topical Application on the Dorsal Skin of Hairless Mice

Alpha-ketoglutarate is a key intermediate in the Krebs cycle, and a rate-limiting cofactor of prolyl-4-hydroxylase. It also has a potent effect on increasing the proline pool during collagen production, but the details underlying the boosting effect on collagen production by alpha-ketoglutarate remain as yet unreported. To investigate the effects of alpha-ketoglutarate on procollagen production and wrinkle formation, we conducted experiments in cultured human dermal fibroblasts and UVB-irradiated hairless mice. Based on ELISA measurements, alpha-ketoglutarate (10 microM) stimulated procollagen production in fibroblasts by 25.6+/-4.6% compared to vehicle (dH(2)O)-treated control cells. Also, we demonstrated that alpha-ketoglutarate increased activities of prolidase, which is known to play an important role in collagen metabolism, in fibroblasts and N-benzyloxycarbonyl-L-proline (Cbz-Pro), prolidase inhibitor, inhibited procollagen synthesis by alpha-ketoglutarate in fibroblasts. To determine the effect of topically applied alpha-ketoglutarate on wrinkle formation, alpha-ketoglutarate (1%) and vehicle (70% propylene glycol, 30% ethanol) were applied on the dorsal skin of UVB-induced hairless mice for twelve weeks. We found that alpha-ketoglutarate decreased wrinkle formation upon long-term topical application. These results suggest that alpha-ketoglutarate diminishes UVB-induced wrinkle formation by increasing collagen production, through a pathway that involves prolidase activation. Therefore, application of alpha-ketoglutarate may represent an effective anti-wrinkle agent for the cosmetic field.

Molecular Markers in Patients with Chronic Wounds to Guide Surgical Debridement

Chronic wounds, such as venous ulcers, are characterized by physiological impairments manifested by delays in healing, resulting in severe morbidity. Surgical debridement is routinely performed on chronic wounds because it stimulates healing. However, procedures are repeated many times on the same patient because, in contrast to tumor excision, there are no objective biological/molecular markers to guide the extent of debridement. To develop bioassays that can potentially guide surgical debridement, we assessed the pathogenesis of the patients' wound tissue before and after wound debridement. We obtained biopsies from three patients at two locations, the nonhealing edge (prior to debridement) and the adjacent, nonulcerated skin of the venous ulcers (post debridement), and evaluated their histology, biological response to wounding (migration) and gene expression profile. We found that biopsies from the nonhealing edges exhibit distinct pathogenic morphology (hyperproliferative/hyperkeratotic epidermis; dermal fibrosis; increased procollagen synthesis). Fibroblasts deriving from this location exhibit impaired migration in comparison to the cells from adjacent nonulcerated biopsies, which exhibit normalization of morphology and normal migration capacity. The nonhealing edges have a specific, identifiable, and reproducible gene expression profile. The adjacent nonulcerated biopsies have their own distinctive reproducible gene expression profile, signifying that particular wound areas can be identified by gene expression profiling. We conclude that chronic ulcers contain distinct subpopulations of cells with different capacity to heal and that gene expression profiling can be utilized to identify them. In the future, molecular markers will be developed to identify the nonimpaired tissue, thereby making surgical debridement more accurate and more efficacious.

An inhibitor of free radical processes decreases the rate of protein synthesis in gunshot wound tissues and decelerates the general adaptation syndrome

The dynamics of total protein and procollagen biosynthesis was studied in the muscle tissue in gunshot wound during the natural healing and after treatment with antioxidant (AO) 2,6-di-tert-butyl-4-methylphenol (BHT). AO treatment considerably decreased the rate of both total protein and procollagen synthesis in the muscle on the third day of healing. BHT notably decreased protein biosynthesis and inhibited corticosteroid production as measured from the electron spin resonance (ESR) signal of reduced adrenodoxin. AO treatment also decreased the rate of protein biosynthesis in bone marrow, thymus, and spleen. The decreased functional activity of the endocrine and immune systems suggests that AO considerably decelerates the systemic body response (general adaptation syndrome) to acute wound. Lipid peroxide radicals and the products of lipid hydroperoxide degradation are proposed as the primary mediators of the body response to stress.

Smooth Muscle α-Actin Deficiency in Myofibroblasts Leads to Enhanced Renal Tissue Fibrosis

Myofibroblasts are a major source of proinflammatory cytokines and extracellular matrix in progressive tissue fibrosis leading to chronic organ failure. Myofibroblasts are characterized by de novo expression of smooth muscle alpha-actin (SMalphaA), which correlates with the extent of disease progression, although their exact role is unknown. In vitro cultured myofibroblasts from kidney of SMalphaA knock-out mice demonstrate significantly more prominent cell motility, proliferation, and type-I procollagen expression than those of wild-type myofibroblasts. These pro-fibrotic properties are suppressed by adenovirus-mediated SMalphaA re-expression, accompanied by down-regulation of focal adhesion proteins. In interstitial fibrosis model, tissue fibrosis area, proliferating interstitial cell number, and type-I procollagen expression are enhanced under SMalphaA deficiency. In mesangioproliferative glomerulonephritis model, cell proliferation in the mesangial area is also enhanced in SMalphaA knock-out mice. Adenoviral SMalphaA introduction into renal interstitium obviously ameliorates tissue fibrosis in interstitial fibrosis model. These results indicate that SMalphaA suppresses the pro-fibrotic properties of myofibroblasts, highlighting the significance of smooth muscle-related proteins in moderating chronic organ fibrosis under pathological conditions.

Regulation of type I procollagen and MMP-1 expression after single or repeated exposure to infrared radiation in human skin

Human skin is daily exposed to infrared (IR) radiation from natural sunlight. However, the effects of IR irradiation on collagen metabolism have not been investigated in human skin in vivo. Here, we examined whether single or repeated (three times a week for 4 weeks) exposure to IR irradiation changes the expressions of type I procollagen and interstitial collagenase (MMP-1). By using immunostaining, Western blotting, and semi-quantitative RT-PCR, we analyzed the protein and mRNA levels of type I procollagen and MMP-1 in young buttock skin. A single dose of IR to human skin increased the expression of type I procollagen within 24 h, but did not change the expression of MMP-1. On the other hand, multiple IR doses reduced the expression of type I procollagen and increased the expression of MMP-1. We also found that TGF-βs may mediate type I procollagen synthesis in IR-irradiated human skin. Our results demonstrate that the regulations of the expressions of type I procollagen and MMP-1 differ in acute and chronically IR-irradiated skin. In particular, decreased collagen levels and increased MMP-1 levels in chronic IR-irradiated skin may be associated with connective tissue damage. Thus, we suggest that repeated exposure to IR irradiation might induce premature skin aging (photoaging) in human skin in vivo.

The NH2-terminal Propeptide of Type I Procollagen Acts Intracellularly to Modulate Cell Function

The function of the NH(2)-terminal propeptide of type I procollagen (N-propeptide) is poorly understood. We now show that a recombinant trimeric N-propeptide interacts with transforming growth factor-beta1 and BMP2 and exhibits functional effects in stably transfected cells. The synthesis of N-propeptide by COS-7 cells results in an increase in phosphorylation of Akt and Smad3 and is associated with a marked reduction in type I procollagen synthesis and impairment in adhesion. In C2C12 cells, N-propeptide inhibits the osteoblastic differentiation induced by BMP2. Our data suggest that these effects are mediated by the interaction of N-propeptide with an intracellular receptor in the secretory pathway, because they are not observed when recombinant N-propeptide is added to the culture medium of either COS-7 or C2C12 cells. Both the binding of N-propeptide to cytokines and its functional properties are entirely dependent on the exon 2-encoded globular domain, and a mutation that substitutes a serine for a highly conserved cysteine in exon 2 abolishes its function. Our findings suggest that N-propeptide performs an important feedback regulatory function and provides a rationale for the prominence of a homotrimeric form of type I procollagen (alpha1 trimer) during vertebrate development.