Temozolomide alleviates breast carcinoma via the inhibition of EGFR/ERK/ MMP-1 pathway with induction of apoptotic events.

PURPOSE: To evaluate the chemotherapeutic activity of temozolomide counter to mammary carcinoma. METHODS: In-vitro anticancer activity has been conducted on MCF7 cells, and mammary carcinoma has been induced in Wistar rats by introduction of 7, 12-Dimethylbenz(a)anthracene (DMBA), which was sustained for 24 weeks. Histopathology, immunohistochemistry, cell proliferation study and apoptosis assay via TUNEL method was conducted to evaluate an antineoplastic activity of temozolomide in rat breast tissue. RESULTS: IC50 value of temozolomide in MCF7 cell has been obtained as 103 µM, which demonstrated an initiation of apoptosis. The temozolomide treatment facilitated cell cycle arrest in G2/M and S phase dose dependently. The treatment with temozolomide suggested decrease of the hyperplastic abrasions and renovation of the typical histological features of mammary tissue. Moreover, temozolomide therapy caused the downregulation of epidermal growth factor receptor, extracellular signal-regulated kinase, and metalloproteinase-1 expression and upstream of p53 and caspase-3 proliferation to indicate an initiation of apoptotic events. CONCLUSIONS: The occurrence of mammary carcinoma has been significantly decreased by activation of apoptotic pathway and abrogation of cellular propagation that allowable for developing a suitable mechanistic pathway of temozolomide in order to facilitate chemotherapeutic approach.

Protective role of Caesalpinia sappan extract and its main component brazilin against blue light-induced damage in human fibroblasts.

BACKGROUND: Ultraviolet (UV) radiation is a well-known factor that causes skin aging. Recently, with the development of technology, the skin has been exposed to not only the UV radiation but also the blue light from electronic devices. Blue light is a high-energy visible light that penetrates deep into the dermal layer, producing reactive oxygen species (ROS) and resulting in skin aging. In this study, we searched for candidate materials that can inhibit blue light-induced skin aging and found Caesalpinia sappan extract (CSE) to be effective. METHODS: Human dermal fibroblasts (HDFs) were treated with various concentrations of CSE and brazilin and exposed to blue light. We measured that antioxidant activity, MMP-1 levels using MMP-1 ELISA, changes in collagen type 1, collagen type 3, MMP-1, and MMP-3 mRNA expressions, and ROS generation. RESULTS: We confirmed that CSE has high absorption of blue light and antioxidant activity. Blue light irradiation at 30 J/cm2 decreased the expression of collagen types 1 and 3, increased the expression of matrix metalloproteinase (MMP)-1 and 3, and decreased the production of ROS in human dermal fibroblasts as compared to those of the nonirradiated group. However, pretreatment with CSE protected against the damage caused by the blue light. Brazilin, a major constituent of C. sappan, had high absorbance in the blue light region and antioxidant activities. Pretreatment with brazilin also inhibited the damage caused by the blue light in the cells. CONCLUSION: CSE and brazilin are potential agents for inhibiting skin aging caused by blue light-induced damage.

Clinical, Histological, and Profibrotic Extracellular Matrix Protein Changes in a Model of Tracheal Stenosis Induced by Cervical Tracheal Autotransplantation.

BACKGROUND: Tracheal stenosis (TS) is a complication of prolonged intubation, tracheotomy, and tracheal surgery that compromises the vascular supply. Animal models are essential for studying its pathophysiology and the effect of interventions. OBJECTIVE: To establish a TS model in rats secondary to tracheal autotransplantation with a graft submerged in bleomycin (Atx-Bleo). Additionally, to evaluate the clinical and histological changes, as well as the expression of newly formed collagen (NFC), isoforms of transforming growth factor beta (TGFß), fibronectin (FN), elastin (ELN), integrin ß1 (ITGß1), and matrix metalloproteinase 1 (MMP1) in TS. METHODS: Twenty Wistar rats were divided into three groups: group I (n = 20) control; group II (n = 10) end-to-end anastomosis of the trachea (tracheoplasty); and group III (n = 10) Atx-Bleo. The animals were evaluated clinically, tomographically, macroscopically, morphometrically, and microscopically. NFC deposition, and the expression of profibrotic and antifibrotic proteins were evaluated in tracheal scars. RESULTS: All animals survived the surgical procedure and the study period. Compared with the other study groups, the Atx-Bleo group developed TS and fibrosis, exhibited higher expression of NFC, TGFß1, TGFß2, FN, ELN, and ITGß1, and mild expression of TGFß3 and MMP1 (p < 0.005; analysis of variance, Dunnett and Tukey tests). CONCLUSION: Atx-Bleo in TS model rats produces tomographic and histological changes, and induces the upregulation of profibrotic proteins (TGFß1, TGFß2, collagen, FN, ELN, ITGß1) and downregulation of antifibrotic proteins (TGFß3, MMP1). Therefore, this model may be used to test new pharmacological treatments for reversing or preventing TS, and conduct basic studies regarding its pathophysiology.

Histological response and expression of collagen, metalloproteinases MMP-1 and MMP-9 and tissue inhibitors of metalloproteinases TIMP-1 and TIMP-2 in fetal membranes following open intrauterine surgery: an experimental study.

OBJECTIVE: To characterize aspects of the repair process by evaluating the tissue collagen density, metalloproteinases and tissue inhibitor of matrix metalloproteinases in the fetal membranes following open fetal surgery for myelomeningocele (MMC). DESIGN: Experimental. SETTING: Two Brazilian hospitals in 2013-2014. POPULATION: 30 fetal membranes collected after elective cesarean deliveries, in patients who underwent open fetal surgery for MMC intrauterine repair. METHODS: Regions within and surrounding the scar area and regions distant from the surgical site were evaluated for collagen concentration and expression of MMP-1, MMP-9, TIMP-1 and TIMP-2. RESULTS: Collagen was increased in regions of scar formation (14.4 ± 2.7%) as compared to unaffected regions (8.0 ± 1.9%) (p < .001). The mean score of MMP-9 in the area of both the scar and suture was also increased above that observed in normal regions (p < .05). Conversely, MMP-1 was reduced in the scar when compared to the normal region and the area adjacent to the scar (suture region) (p < .05). TIMP-1 was increased in the suture region compared to the normal region (p < .05) while TIMP-2 was reduced in the scar region when compared to the other two regions (p < .05). The membrane repair process was also influenced by the number of previous pregnancies and gestational age at the time of surgery. CONCLUSION: Reparative activity of the fetal membrane after open fetal surgery involves up-regulation of collagen production and differential involvement of MMPs and TIMPs.

Prolactin Protects the Structural Integrity of Human Fetal Membranes by Downregulating Inflammation-induced Secretion of Matrix Metalloproteinases.

Prolactin (PRL) is a pleiotropic hormone with a key role in pregnancy. In fetal membranes, PRL can regulate the secretion of pro-inflammatory factors, which induces the activation of matrix metalloproteinases (MMPs). The increase and activation of MMPs deregulate the turnover of the extracellular matrix in the fetal membranes, altering its structure and function, causing premature rupture of the membranes and preterm labor. In this work, we evaluate the effect of PRL upon the secretion of MMP-1, MMP-2, MMP-9, MMP-13, and the tissue inhibitors of metalloproteinases (TIMPs) in human fetal membranes after lipopolysaccharide (LPS) challenge. Nine fetal membranes from healthy non-laboring cesarean deliveries at term were cultured in a 2-independent chamber system and pre-treated with 250, 500, 1000 or 4000 ng/ml of PRL for 24 h, then choriodecidual region was stimulated with 500 ng/ml of LPS plus fresh PRL for 24 h. The MMPs and TIMPs secretion were quantified by ELISA, additionally MMP-2 and MMP-9 gelatinolytic activity was measured by zymography. LPS induced the MMP-9 and MMP-1 secretion, but no MMP-2 or MMP-13 in comparison with basal levels. PRL co-treatment decreased the MMP-2, MMP-9 and MMP-1 secretion induced by LPS. The active forms were present in the tissue extract, showing a response consistent with the secretion profile. TIMP-1 and TIMP-2 secretion was decreased after LPS treatment and the PRL co-treatment reverts this effect. The present results support that PRL may favor the balance between these factors involved in the structural maintenance of fetal membranes in an inflammatory event.

Treatment with Hyaluronic Acid and Collagen-Polyvinylpyrrolidone Improves Extracellular Matrix Assembly for Scarring after Tracheal Resection.

Treatment of tracheal stenosis is occasionally performed in combination with wound healing modulators to manipulate new extracellular matrix (ECM) formation and prevent fibrosis. Hyaluronic acid (HA) and collagen-polyvinylpyrrolidone (collagen-PVP) decrease fibrosis in experimental tracheal healing. However, they have not been used clinically as their effect on ECM components, which modify tracheal scarring, has not been described. Objective. To evaluate the effect of the application of HA, collagen-PVP, a mixture of HA and collagen-PVP (HA+collagen-PVP), and mitomycin C on the expression of decorin, matrix metalloproteinase 1 (MMP1), and MMP9, as well as the type of collagen and deposits formed in the scar after resection and end-to-end anastomosis (REEA) of the cervical trachea using an experimental model. Materials and Methods. Thirty dogs underwent REEA of the cervical trachea and were treated with different wound healing modulators: group I (n = 6), control; group II (n = 6), HA; group III (n = 6), collagen-PVP; group IV (n = 6), HA+collagen-PVP; and group V (n = 6), mitomycin C. The dogs were evaluated clinically and endoscopically for 4 weeks. Subsequently, macroscopic and microscopic changes, expression of ECM proteins, and collagen deposition in tracheal scars were analysed. Results. Groups II, III, and IV showed reduced endoscopic, macroscopic, and microscopic inflammation, improved neovascularization, high decorin expression (p < 0.01, analysis of variance (ANOVA)), and moderate expression of MMP1 (p < 0.003, ANOVA) and type I and III collagen (p < 0.05, Kruskal-Wallis). Groups IV and V developed fewer collagen deposits (p < 0.001, ANOVA). Conclusion. Treatment with HA and collagen-PVP improved post-REEA healing by increasing neovascularization, stimulating the expression of decorin, and regulating the expression of MMP1, as well as type I and III collagen and their deposition.

Polysaccharide-rich hydrogel formulation combined with photobiomodulation repairs UV-induced photodamage in mice skin.

Prolonged skin exposure to ultraviolet radiation (UVR) induces premature aging in both the epidermis and the dermis. Chronic exposure to UVR induces the activation of mitogen-activated protein kinase (MAPK) signaling pathway, activating c-Jun, c-Fos expression, and transcription factor of AP-1 activating protein. AP-1 activation results in the positive induction of matrix metalloproteinase (MMP) synthesis, which degrade skin collagen fibers. Polysaccharides from the fruit of Lycium barbarum (LBP fraction) have a range of activities and have been demonstrate to repair the photodamage. In different approaches, laser application aims to recover the aged skin without destroying the epidermis, promoting a modulation, called photobiomodulation (PBM), which leads to protein synthesis and cell proliferation, favoring tissue repair. Here we developed a topical hydrogel formulation from a polysaccharide-rich fraction of Lycium barbarum fruits (LBP). This formulation was associated with PBM (red laser) to evaluate whether the isolated and combined treatments would reduce the UVR-mediated photodamage in mice skin. Hairless mice were photoaged for 6 weeks and then treated singly or in combination with LBP and PBM. Histological, immunohistochemistry, and immunofluorescence analyses were used to investigate the levels of c-Fos, c-Jun, MMP-1, -2, and -9, collagen I, III, and FGF2. The combined regimen inhibited UVR-induced skin thickening, decreased the expression of c-Fos and c-Jun, as well as MMP-1, -2, and -9 and concomitantly increased the levels of collagen I, III, and FGF2. The PBM in combination with LBP treatment is a promising strategy for the repair of photodamaged skin, presenting potential clinical application in skin rejuvenation.

Association of the polymorphisms of the genes APOC3 (rs2854116), ESR2 (rs3020450), HFE (rs1799945), MMP1 (rs1799750) and PPARG (rs1801282) with lipodystrophy in people living with HIV on antiretroviral therapy: a systematic review.

The aim of this study was to perform a systematic review to identify data reported in the literature concerning the association of APOC3 (rs2854116), ESR2 (rs3020450), HFE (rs1799945), MMP1 (rs1799750) and PPARG (rs1801282) polymorphisms with lipodystrophy in people living with HIV (PLWHIV) on antirretroviral therapy. The research was conducted in six databases and the studies were selected in two steps. First, a search was undertaken in the following electronic databases: PubMed, Science Direct, Medline, World Wide Science, Directory of Open Access Journals, Scielo, Lilacs and Medcarib. The titles and abstracts of 24,859 articles were read to select those that match the elegibilty criteria. Five papers that addressed the association of HAART, lipodystrophy and polymorphisms were selected for the review. There was no association between the polymorphisms of the genes APOC3 and PPARG and lipodystrophy. Another study described an association between the variant allele (G) of HFE and protection concerning the development of lipoatrophy (0.02) when compared with the reference allele (C). On the other hand, the variant allele (T) of the ESR2 gene was associated with the development of lipoatrophy (p = 0.007) when compared with the reference allele (C). In addition, the genotype and the variant allele of the gene MMP1 (2G) were associated with lipodystrophy in PLWHIV on HAART (p = 0.0002 and p = 0.0008, respectively). Therefore, further studies with other populations, involving PLWHIV on HAART are necessary to better understand the role of genetic markers, which may be involved in a predisposition to lipodystrophy.

Abilities of protocatechuic acid and its alkyl esters, ethyl and heptyl protocatechuates, to counteract UVB-induced oxidative injuries and photoaging in fibroblasts L929 cell line.

Ultraviolet B (UVB) radiation triggers the activation of many reactive oxygen species (ROS)-sensitive signaling pathways, resulting in the induction of skin damage that can progress to premature skin aging with long-term exposure. Even after the cessation of UVB radiation, the activated photosensitizers can still cause cellular injury. Thus, the use of photoprotectors that inhibit or prevent intracellular ROS production during or after UV exposure is one alternative to counteract UV-induced oxidative damage. The present study investigated the photoprotective activity of protocatechuic acid (P0) and its alkyl esters ethyl protocatechuate (P2) and heptyl protocatechuate (P7) against UVB-induced damage in L929 fibroblasts by evaluating biomarkers of oxidative stress and photoaging. P0, P2 and P7 markedly increased cell viability after UVB exposure. This protective effect was related to the ability of these compounds to absorb UVB and restore cellular redox balance even 24 h after UVB exposure. P0, P2 and P7 also decreased oxidative damage to membrane lipids, mitochondrial membrane potential, and DNA. They also inhibited the nuclear translocation of NF-κB p65 and downregulated the expression of the photoaging-related proteins matrix metalloproteinases-1 and -9 and cyclooxygenase-2. As the lipophilicity of the P0 derivatives increased, their antioxidant potency increased, but more pronounced cytotoxic effects were also detected. In summary, P0 and P2 may be promising candidates for the prevention and treatment of UVB-induced skin photodamage and photoaging.

Effects of the serine protease inhibitor rBmTI-A in an experimental mouse model of chronic allergic pulmonary inflammation.

To evaluate whether a recombinant serine protease inhibitor (rBmTI-A) modulates inflammation in an experimental model of chronic allergic lung inflammation. Balb/c mice were divided into four groups: SAL (saline), OVA (sensitized with ovalbumin), SAL + rBmTI-A (control treated with rBmTI-A) and OVA + rBmTI-A (sensitized with ovalbumin and treated with rBmTI-A). The animals received an intraperitoneal injection of saline or ovalbumin, according to the group. The groups received inhalation with saline or ovalbumin and were treated with rBmTI-A or saline by nasal instillation. After 29 days, we evaluated the respiratory mechanics; bronchoalveolar lavage fluid (BALF); cytokines; MMP-9, TIMP-1; eosinophils; collagen and elastic fibre expression in the airways; and the trypsin-like, MMP-1, and MMP-9 lung tissue proteolytic activity. Treatment with rBmTI-A reduced the trypsin-like proteolytic activity, the elastance and resistance maximum response, the polymorphonuclear cells, IL-5, IL-10, IL-13 and IL-17A in the BALF, the expression of IL-5, IL-13, IL-17, CD4+, MMP-9, TIMP-1, eosinophils, collagen and elastic fibres in the airways of the OVA + rBmTI-A group compared to the OVA group (p < 0.05). rBmTI-A attenuated bronchial hyperresponsiveness, inflammation and remodelling in this experimental model of chronic allergic pulmonary inflammation. This inhibitor may serve as a potential therapeutic tool for asthma treatment.

Photobiomodulation of Matrix Metalloproteinases in Rat Calcaneal Tendons.

Objective: The main objective was to verify the modulatory effects of MMP-1, MMP-3, and MMP-13 levels on the partially injured calcaneal tendons of rat exposure to photobiomodulation. Background: Photobiomodulation has been shown to have anti-inflammatory and regenerative effects on tendon injuries. However, there is still uncertainty regarding the beneficial effects in matrix metalloproteinase (MMP) levels, especially MMP-1, -3, and -13. Materials and methods: Sixty-five male Wistar rats were used. Sixty were submitted to a direct trauma on the calcaneal tendons and were randomly distributed into the following six groups: LASER 1, 3, and 7 (10 partially injured calcaneal tendons in each group treated with photobiomodulation for 1, 3, and 7 days, respectively) and Sham 1, 3, and 7 (same injury, with simulated photobiomodulation). The remaining five animals were allocated to the normal group (no injury or treatment procedure). The 780 nm low-level laser was applied with 70 mW of mean power and 17.5 J/cm2 of fluency for 10 sec, once a day. The tendons were surgically removed and analyzed for MMP-1, MMP-3, and MMP-13 through immunohistochemistry. Results: MMP-3 levels remained close to normal in all experimental groups (p > 0.05); however, reductions (p < 0.05) in MMP-1 and MMP-13 levels were detected in the groups submitted to one, three, and seven low level laser therapy applications. Conclusions: The photobiomodulation protocol was able to reduce MMP-1 and MMP-13 levels in injured calcaneal tendons.

Matrix Metalloproteinases System and Types of Fibrosis in Rat Heart during Late Pregnancy and Postpartum.

Background and objectives: Cardiac remodeling in pregnancy and postpartum is poorly understood. The aim of this study was to evaluate changes in cardiac fibrosis (pericardial, perivascular, and interstitial), as well as the expression of matrix metalloproteinases (MMP-1, MMP-2, and MMP-9) and their inhibitors (Tissue inhibitors of metalloproteinases, TIMP-1 and TIMP-4) during late pregnancy and postpartum in rat left ventricle. Materials and Methods: Female Sprague-Dawley rats were used for this study. Rats were divided three groups: non-pregnant, late pregnancy, and postpartum. The heart was weighed and cardiac fibrosis was studied by conventional histological procedures. The expression and transcript level of target proteins were evaluated using immunoblot techniques and quantitative PCR. Results: The experiments showed an increase of perivascular, pericardial, and interstitial fibrosis in heart during pregnancy and its reversion in postpartum. Moreover, in late pregnancy, MMP-1, MMP-2, and MMP-9 metalloproteinases were downregulated and TIMP-1 and TIMP-4 were upregulated in left ventricle. Conclusions: Our data suggest that the metalloproteinases system is involved in the cardiac extracellular matrix remodeling during pregnancy and its reversion in postpartum, this improves the knowledge of the adaptive cardiac remodeling in response to a blood volume overload present during pregnancy.

Dihydrocaffeic Acid Prevents UVB-Induced Oxidative Stress Leading to the Inhibition of Apoptosis and MMP-1 Expression via p38 Signaling Pathway.

Chronic UVB exposure promotes oxidative stress, directly causes molecular damage, and induces aging-related signal transduction, leading to skin photoaging. Dihydrocaffeic acid (DHCA) is a phenolic compound with potential antioxidant capacity and is thus a promising compound for the prevention of UVB-induced skin photodamage. The aim of this study was to evaluate the antioxidant and protective effect of DHCA against oxidative stress, apoptosis, and matrix metalloproteinase (MMP) expression via the mitogen-activated protein kinase (MAPK) signaling pathway on L929 fibroblasts irradiated with UVB. DHCA exhibited high antioxidant capacity on 2,2-diphenyl-1-picrylhydrazyl (DPPH•), 2,2-azinobis-3-ethylbenzothiazoline-6-sulphonic acid (ABTS•+), and xanthine/luminol/xanthine oxidase (XOD) assays and reduced UVB-induced cell death in the neutral red assay. DHCA also modulated oxidative stress by decreasing intracellular reactive oxygen species (ROS) and extracellular hydrogen peroxide (H2O2) production, enhancing catalase (CAT) and superoxide dismutase (SOD) activities and reduced glutathione (GSH) levels. Hence, cellular damage was attenuated by DHCA, including lipid peroxidation, apoptosis/necrosis and its markers (loss of mitochondria membrane potential, DNA condensation, and cleaved caspase 9 expression), and MMP-1 expression. Furthermore, DHCA reduced the phosphorylation of MAPK p38. These findings suggest that DHCA can be used in the development of skin care products to prevent UVB-induced skin damage.

Short-term Changes After Corticosteroid Injections Into the Normal Tendons of Rabbits: A Controlled Randomized Study.

BACKGROUND: Corticosteroid injections in or around tendons for the treatment of athletic injuries are a common practice among orthopaedic surgeons and are apparently efficacious in the short term, although controversies persist related to local complications. PURPOSE: This study evaluated short-term (48 hours) biomechanical, biochemical, and histological alterations after a single injection of betamethasone into the normal tendons of rabbits. STUDY DESIGN: Controlled laboratory study. METHODS: A total of 72 New Zealand White rabbits were randomly divided into 2 groups: the test group-in which 36 animals underwent 1 intratendinous injection of betamethasone (1.4 mg / 0.2 mL) in the right calcaneal tendon; the control group-in which the right calcaneal tendon of 36 animals was injected with saline (placebo control group) and the left calcaneal tendon was left untreated for normal standards (normal control). Forty-eight hours later, animals were euthanized and tendons were harvested. Metalloproteinase (MMP1 and MMP2) and interleukin (IL1 and IL6) expression levels, biomechanical resistance (load × elongation parameters), and histomorphometry (hematoxylin and eosin and picrosirius red stains for collagen fibers, tenocytes, and inflammatory cells) were analyzed in the tendons. RESULTS: The test group had a significant reduction in MMP2 expression as compared with the control groups ( P = .027). Regarding the other parameters, there were no additional significant differences between the groups. CONCLUSION: A single injection of corticosteroid into normal calcaneal tendons did not trigger acute local morphological, structural, or biomechanical injuries at 48 hours, but it did promote a significant decrease in MMP2 levels. Additional studies are needed with increased duration of follow-up, various doses, and multiple injections and in tendinopathic models. CLINICAL RELEVANCE: Some previous studies demonstrated early structural changes in tendons after a single corticosteroid injection, which was not corroborated by the present study. Metalloproteinase decrease is usually associated with a reduction in collagen degradation, which would be protective for the healing process. More studies are necessary to confirm the possible beneficial effect of these results in the long term and for tendinopathies.

R-Spondin-2 Is Upregulated in Idiopathic Pulmonary Fibrosis and Affects Fibroblast Behavior.

Idiopathic pulmonary fibrosis (IPF) is characterized by the expansion of the myofibroblast population, excessive extracellular matrix accumulation, and destruction of the lung parenchyma. The R-spondin family (RSPO) comprises a group of proteins essential for development. Among them, RSPO2 is expressed primarily in the lungs, and its mutations cause severe defects in the respiratory tract. Interestingly, RSPO2 participates in the canonical Wingless/int1 pathway, a critical route in the pathogenesis of IPF. Thus, the aim of this study was to examine the expression and putative role of RSPO2 in this disease. We found that RSPO2 and its receptor leucine-rich G protein-coupled receptor 6 were upregulated in IPF lungs, where they localized primarily in fibroblasts and epithelial cells. Stimulation of IPF and normal lung fibroblasts with recombinant human RSPO2 resulted in the deregulation of numerous genes, although the transcriptional response was essentially distinct. In IPF fibroblasts, RSPO2 stimulation induced the up- or downregulation of several genes involved in the Wingless/int1 pathway (mainly from noncanonical signaling). In both normal and IPF fibroblasts, RSPO2 modifies the expression of genes implicated in several pathways, including the cell cycle and apoptosis. In accordance with gene expression, the stimulation of normal and IPF fibroblasts with RSPO2 significantly reduced cell proliferation and induced cell death. RSPO2 also inhibited collagen production and increased the expression of matrix metalloproteinase 1. Silencing RSPO2 with shRNA induced the opposite effects. Our findings demonstrate, for the first time to our knowledge, that RSPO2 is upregulated in IPF, where it appears to have an antifibrotic role.

Collagen peptides modulate the metabolism of extracellular matrix by human dermal fibroblasts derived from sun-protected and sun-exposed body sites.

Clinical data published in recent years have demonstrated positive effects of collagen hydrolysate (CH) on skin aging clinical signs. CH use as food supplement has a long history; however, few studies have addressed the underlying purpose of CH on the cellular and molecular biology of skin cells that could elucidate clinical improvement findings. Wide diversity of characteristics has been reported for dermal fibroblasts derived from different body sites and it is unknown whether collagen peptides could modulate differently cells from chronological aged and photoaged skin areas. This study investigated the influence of CH on the extracellular matrix metabolism and proliferation of human dermal fibroblasts (HDFs) derived from chronological aged (sun-protected) and photoaged (sun-exposed) body sites. CH treatment did not affect cellular proliferation of either cell cultures, but notably modulated cell metabolism in monolayer model, increasing the content of dermal matrix precursor and main protein, procollagen I and collagen I, respectively. These effects were confirmed in the human dermal equivalent model. The increase in collagen content in the cultures was attributed to stimulation of biosynthesis and decreased collagen I metabolism through inhibition of metalloproteinase activity (MMP) 1 and 2. Modulation of CH in dermal metabolism did not differ between cells derived from sun-protected and sun-exposed areas, although lower concentrations of CH seemed to be enough to stimulate sun-exposed-derived HDFs, suggesting more pronounced effect in these cells. This study contributes to understanding the biological effects of CH on skin cells and viability of its use as a functional ingredient in food supplements.

The Role of Proteinase-Activated Receptors 1 and 2 in the Regulation of Periodontal Tissue Metabolism and Disease.

Proteinase-activated receptors 1 (PAR1) and 2 (PAR2) are the most highly expressed members of the PAR family in the periodontium. These receptors regulate periodontal inflammatory and repair processes through their activation by endogenous and bacterial enzymes. PAR1 is expressed by the periodontal cells such as human gingival fibroblasts, gingival epithelial cells, periodontal ligament cells, osteoblasts, and monocytic cells and can be activated by thrombin, matrix metalloproteinase 1 (MMP-1), MMP-13, fibrin, and gingipains from Porphyromonas gingivalis. PAR2 is expressed by neutrophils, osteoblasts, oral epithelial cells, and human gingival fibroblasts, and its possible activators in the periodontium are gingipains, neutrophil proteinase 3, and mast cell tryptase. The mechanisms through which PARs can respond to periodontal enzymes and result in appropriate immune responses have until recently been poorly understood. This review discusses recent findings that are beginning to identify a cardinal role for PAR1 and PAR2 on periodontal tissue metabolism.

The effect of potassium diclofenac and dexamethasone on MMP-1 gene transcript levels during experimental tooth movement in rats.

OBJECTIVES: The aim of this study was to analyze MMP-1 transcript levels in periodontal tissues of rats that underwent orthodontic treatment using potassium diclofenac and dexamethasone at different stages of tooth movement. SETTING AND SAMPLE POPULATION: The sample comprised of ninety male Wistar rats. MATERIAL AND METHODS: A closed nickel-titanium coil spring was used to apply a force of 50 cN to move the maxillary right first molars mesially. One group received daily doses of 0.9% saline solution, the second group received daily doses of 5 mg/kg potassium diclofenac, and the third group received daily doses of 0.5 mg/kg dexamethasone. Tooth movement was observed on days 0, 1, 3, 7, and 14. MMP-1 transcript levels were evaluated by real-time polymerase chain reaction and the results were compared between groups by three-way ANOVA, with a significance level of 0.05. RESULTS: Transcript levels increased in groups that received the coil spring treatment on all days of the experiment. MMP-1 expression was found to be decreased in groups treated with potassium diclofenac and dexamethasone compared to that in the control group, on days 1, 3, 5, and 7. CONCLUSIONS: The application of orthodontic forces significantly increased MMP-1 transcript levels. The use of anti-inflammatory drugs may have an inhibitory effect on MMP-1 expression.

Cimetidine Reduces Interleukin-6, Matrix Metalloproteinases-1 and -9 Immunoexpression in the Gingival Mucosa of Rat Molars With Induced Periodontal Disease.

BACKGROUND: Histamine seems to act, via H2 receptor, on inflammatory processes by stimulating interleukin (IL)-6 and matrix metalloproteinase (MMP) release. As cimetidine is an H2 receptor antagonist, the authors hypothesize that this antiulcer drug reduces IL-6, MMP-1, and MMP-9 immunoexpression in gingiva with induced periodontal disease (PD). To confirm a possible modulatory role of IL-6 on MMPs, the relationship between IL-6/MMP-1 and IL-6/MMP-9 immunoexpression was evaluated. METHODS: Forty-six male rats were distributed into the cimetidine group (CimG: received daily intraperitoneal injections of 100 mg/kg of body weight of cimetidine) or the saline group (SG). PD was induced by cotton ligature around the maxillary left first molars (PDSG and PDCimG). The right molars were used as controls (SG and CimG). After 7, 15, 30, and 50 days, maxillary fragments were processed for paraffin embedding or for transmission electron microscopy. Tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts in the alveolar process surface and number of IL-6, MMP-1, and MMP-9-immunolabeled cells in the gingival mucosa were quantified. Statistical analyses were performed (P ≤0.05). RESULTS: In PDSG and PDCimG, gingival mucosa exhibited few collagen fibers among numerous inflammatory cells. In PDCimG, the number of TRAP-positive osteoclasts and IL-6, MMP-1, and MMP-9-immunolabeled cells was significantly lower than in PDSG at all periods. A positive correlation between IL-6/MMP-1 and IL-6/MMP-9 was detected in PDSG and PDCimG. CONCLUSION: Cimetidine decreases bone loss through reduction of osteoclast number and induces reduction of IL-6, MMP-1, and MMP-9 immunoexpression, reinforcing the idea that the beneficial effect of cimetidine in PD may be due to reduction of IL-6 immunolabeling in the inflamed gingival mucosa.

A three-dimensional cell culture system as an in vitro canine mammary carcinoma model for the expression of connective tissue modulators.

In this study, derived complex carcinoma (CC) and simple carcinoma (SC) cell lines were established and cultured under two-dimensional (2D) and three-dimensional (3D) conditions. The 3D was performed in six-well AlgiMatrix™ (LifeTechnologies®, Carlsbad, CA, USA) scaffolds, resulting in spheroids sized 50-125 µm for CC and 175-200 µm for SC. Cell viability was demonstrated up to 14 days for both models. Epidermal growth factor receptor (EGFR) was expressed in CC and SC in both systems. However, higher mRNA and protein levels were observed in SC 2D and 3D systems when compared with CC (P < 0.005). The connective tissue modulators, metalloproteinases-1, -2, -9 and -13 (MMPs), relaxin receptors 1 and 2 (RXR1 and RXR2) and E-cadherin (CDH1) were quantitated. All were upregulated similarly when canine mammary tumour (CMT)-derived cell lines were cultured under 3D AlgiMatrix, except CDH1 that was downregulated (P < 0.005). These results are promising towards the used of 3D system to increase a high throughput in vitro canine tumour model.