Correlation of serum oestrogen with salivary calcium in post-menopausal women with and without oral dryness feeling

doi: 10.1111/j.1741‐2358.2011.00580.x
Correlation of serum oestrogen with salivary calcium in post‐menopausal women with and without oral dryness feeling Objective: The aim of this study was to verify the association between salivary calcium and serum oestrogen levels with oral dryness in post‐menopausal women. Also, the correlation between these variables was evaluated. Methods: A case‐control study was carried out on 60 selected menopausal women with and without oral dryness feeling (30 as case and 30 as control) conducted in the Department of Oral Medicine and Radiology, Maharishi Markandeshar University, Mullana, India. Paraffin‐stimulated saliva samples were obtained by expectoration. Salivary calcium concentrations were assessed colorimetrically using Arsenazo III reaction. The serum oestrogen concentration was measured using ELISA. Statistical analysis of Student’s t‐test and Pearson correlation was used. Results: There was significant difference in mean values of both salivary calcium concentration and serum oestrogen between case and control groups. The result obtained also showed that an inverse correlation was found between salivary calcium concentration and serum oestrogen levels in both the groups and in total sample size. Conclusion: Oral dryness in post‐menopausal women is associated with high levels of salivary calcium and low levels of serum oestrogen. The concentrations of salivary calcium and serum oestrogen are inversely correlated in post‐menopausal women, regardless of the presence or absence of oral dryness.     

Region‐specific effects of oestradiol on adipose‐derived stem cell differentiation in post‐menopausal women

The goal of this study was to determine the effect of acute transdermal 17β‐oestradiol (E₂) on the adipogenic potential of subcutaneous adipose‐derived stem cells (ASC) in post‐menopausal women. Post‐menopausal women (n = 11; mean age 57 ± 4.5 years) were treated for 2 weeks, in a randomized, cross‐over design, with transdermal E₂ (0.15 mg) or placebo patches. Biopsies of abdominal (AB) and femoral (FEM) subcutaneous adipose tissue (SAT) were obtained after each treatment and mature adipocytes were analysed for cell size and ASC for their capacity for proliferation (growth rate), differentiation (triglyceride accumulation) and susceptibility to tumour necrosis factor alpha‐induced apoptosis. Gene expression of oestrogen receptors α and β (ESR1 and ESR2), perilipin 1 and hormone‐sensitive lipase (HSL), was also assessed. In FEM SAT, but not AB SAT, 2 weeks of E₂ significantly (P = 0.03) increased ASC differentiation and whole SAT HSL mRNA expression (P = 0.03) compared to placebo. These changes were not associated with mRNA expression of oestrogen receptors α and β, but HSL expression was significantly increased in FEM SAT with transdermal E₂ treatment. Adipose‐derived stem cells proliferation and apoptosis did not change in either SAT depot after E₂ compared with placebo. Short‐term E₂ appeared to increase the adipogenic potential of FEM, but not AB, SAT in post‐menopausal women with possible implications for metabolic disease. Future studies are needed to determine longer term impact of E₂ on regional SAT accumulation in the context of positive energy imbalance.     

Smad‐induced alterations of matrix metabolism by a myristoyl tetra peptide

Regulation of extracellular matrix (ECM) components is essential for tissue homeostasis and function. We screened a small peptide that induces ECM protein synthesis for its usefulness in protecting keratinocytes. In this report, we demonstrate that myristoyl tetrapeptide Ala‐Ala‐Pro‐Val (mAAPV) stimulates the expression of ECM proteins and inhibits the expression of metalloproteinases (MMPs) that degrade ECM proteins in Hs68 human fibroblast cells. In order to elucidate the underlying molecular mechanisms for the effects of mAAVP, we investigated the changes in gene expression in the presence of mAAPV using a cDNA microarray. Treatment with mAAPV resulted in decreased expression of MMP‐related genes such as MMP1, MMP3, TIMP1 and TIMP3 and increased expression of collagen genes, including COL1A1, COL1A2, COL3A1, COL5A1 and COL6A3. The pattern of gene expression regulated by mAAPV was very similar to that of gene expression induced by transforming growth factor (TGF)‐β, indicating that the TGF‐β signaling pathway is crucial for simultaneous activation of several ECM‐related genes by mAAPV. We examined whether the activation of SMAD, a downstream protein of TGF‐β receptor, is involved in the signal transduction pathway induced by mAAPV. The results demonstrate that mAAVP directly activates SMAD2 and induces SMAD3 to bind to DNA. In conclusion, our results demonstrate that mAAPV both enhances the expression of collagen and inhibits its degradation via production of protease inhibitors that prevent enzymatic breakdown of the ECM. The results suggest that mAAPV would be a useful ECM‐protecting agent. Copyright © 2014 John Wiley & Sons, Ltd.     

Absence of tissue inhibitor of metalloproteinases 3 disrupts alveologenesis in the mouse

Tissue inhibitors of metalloproteinases (TIMPs) regulate extracellular matrix (ECM) degradation by matrix metalloproteinases (MMPs) throughout lung development. We examined lungs from TIMP3 null mice and found significant air space enlargement compared with wild type (WT) animals during a time course spanning early alveologenesis (post‐partum days 1, 5, 9 and 14). Trichrome staining revealed a similar pattern of collagen distribution in the walls of nascent alveoli; however, the alveolar walls of TIMP3 mutant mice appeared to be thinner than controls. Assessment of MMP2 and MMP9 activities by gelatin zymography demonstrated a significant elevation in the active form of MMP2 at post‐partum days 1 and 5. Treatment of null pregnant dams with a broad spectrum synthetic metalloproteinase inhibitor, GM6001, on embryonic day 16.5 enhanced the formation of primitive alveoli during the saccular stage of lung development as evidenced by a partial, but significant, rescue of alveolar size in post‐partum day 1 animals. We propose that increased MMP activity in the absence of TIMP3 enhances ECM proteolysis, upsetting proper formation of primitive alveolar septa during the saccular stage of alveologenesis. Therefore, TIMP3 indirectly regulates alveolar formation in the mouse. To our knowledge, ours is the first study to demonstrate that in utero manipulation of the TIMP/MMP proteolytic axis, to specifically inhibit proteolysis, significantly affects lung development.     

Topical KGF treatment as a therapeutic strategy for vaginal atrophy in a model of ovariectomized mice

One of the most frequent complaints for post‐menopausal women is vaginal atrophy, because of reduction in circulating oestrogens. Treatments based on local oestrogen administration have been questioned as topic oestrogens can reach the bloodstream, thus leading to consider their safety as controversial, especially for patients with a history of breast or endometrial cancers. Recently, growth factors have been shown to interact with the oestrogen pathway, but the mechanisms still need to be fully clarified. In this study, we investigated the effect of keratinocyte growth factor (KGF), a known mitogen for epithelial cells, on human vaginal mucosa cells, and its potential crosstalk with oestrogen pathways. We also tested the in vivo efficacy of KGF local administration on vaginal atrophy in a murine model. We demonstrated that KGF is able to induce proliferation of vaginal mucosa, and we gained insight on its mechanism of action by highlighting its contribution to switch ERα signalling towards non‐genomic pathway. Moreover, we demonstrated that KGF restores vaginal trophism in vivo similarly to intravaginal oestrogenic preparations, without systemic effects. Therefore, we suggest a possible alternative therapy for vaginal atrophy devoid of the risks related to oestrogen‐based treatments, and a patent (no. RM2012A000404) has been applied for this study.     

Photobiomodulation alters matrix protein activity in stressed fibroblast cells in vitro

A balance is maintained between matrix synthesis and degradation, and a prolonged increase in matrix metalloproteinases (MMPs) affects healing. Photobiomodulation (PBM) speeds up healing and alters wound environment. The study aimed to determine changes in protein and gene expression of collagen type 1 (Col‐I), MMP‐3 and ‐9 and TIMP‐1 in fibroblasts irradiated at 660 or 830 nm. Commercially purchased human skin fibroblast cells were modeled into five groups namely, normal, normal wounded, diabetic wounded, hypoxic wounded and diabetic hypoxic wounded. Control cells were sham irradiated. Laser irradiation was conducted at 660 or 830 nm (108/or 94 mW, 9.1 cm², 420/or 483 s) with 5 J/cm². Forty‐eight hours post‐irradiation, protein expression of TIMP‐1, MMP‐3, ?9 and Col‐I was determined by flow cytometry and immunofluorescence, and gene expression by real‐time RT‐PCR. There was an increase in TIMP‐1 and Col‐I, and a decrease in MMP‐3 and ‐9, as well as an alteration in mRNA expression of MMP3, MMP9, TIMP1 and COL1A1 in irradiated cells. Due to the responsiveness of the diabetic hypoxic wounded model, the findings propose this model as appropriate for wound healing studies and suggest that PBM promotes the remodeling phase of wound healing by decreasing matrix degradation and upregulating synthesis.      

Gonadal steroids regulate the expression of aggrecanases in human endometrial stromal cells in vitro

The human endometrium undergoes cyclic change during each menstrual cycle in response to gonadal steroids. Proteolysis of endometrial extracellular matrix (ECM) is necessary to prepare this dynamic tissue for pregnancy. Proteolytic enzymes such as matrix metalloproteinase (MMP) and closely related a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS) have been assigned key roles in the highly regulated cyclic remodelling of the endometrial ECM. We have previously shown that ADAMTS‐1 undergoes spatiotemporal changes in human endometrial stromal cells under the regulation of gonadal steroids. This suggests that other ADAMTS subtypes, known as aggrecanases, may contribute to the ECM remodelling events that occur in female physiological cycles and in preparation for pregnancy. To determine whether progesterone (P4), 17β‐estradiol (E2), or dihydrotestosterone (DHT), alone or in combination, are capable of regulating ADAMTS‐4, ‐5, ‐8 or ‐9 expression in human endometrial stromal cells in vitro. Real‐time quantitative PCR and Western blot analysis were used to measure ADAMTSs mRNA and protein levels in primary cultures of human endometrial stromal cells (n = 12). P4, DHT but not E2 have regulatory effects on ADAMTS‐8, ‐9 and ‐5 expression. Combined treatment with gonadal steroids did not show any synergistic or antagonistic effects. However, the synthetic steroid antagonists RU486 and hydroxyflutamide specifically inhibited the P4‐ or DHT‐mediated regulatory effects on ADAMTS expression. These studies provide evidence that the regulation of aggrecanases by gonadal steroids in human endometrial stromal cells may play an important role during decidualization.     

Evaluation of Ki‐67 and Bcl‐2 antigen expression in breast carcinomas of women treated with raloxifene

Objectives: To evaluate the effect of raloxifene on Ki‐67 and Bcl‐2 antigen expression in operable, stage II, oestrogen‐receptor‐positive invasive ductal breast carcinomas. Materials and methods: Twenty post‐menopausal women who had taken 60 mg of raloxifene daily for 28 days prior to definitive surgery were enrolled in the investigation. Two tumour samples were obtained by incisional biopsy during the study, one at the time of confirmation of diagnosis of invasive ductal carcinoma and evaluation of oestrogen receptor status, and the other 29 days later, at the time of definitive surgery. Immunohistochemistry was performed on tumour samples, prior to and after raloxifene treatment, to evaluate Ki‐67 and Bcl‐2 expression. Friedman and McNemar tests were used for statistical analysis of the data, significance being established at 5%. Results: Mean percentage of Ki‐67‐stained nuclei was 24.86 ± 2.95 prior to raloxifene treatment and 13.33 ± 1.52 after treatment (P < 0.001). Prior to raloxifene treatment, only 9/20 cases (45%) were classified as Bcl‐2‐positive, whereas after treatment, 17/20 (85%) were classified as Bcl‐2‐positive (P < 0.013). Conclusions: Raloxifene treatment significantly reduced Ki‐67 antigen expression and increased Bcl‐2 expression in breast carcinomas of post‐menopausal women.     

四周尾部悬吊大鼠颈总动脉MMP9和TIMP1的表达及活性改变

目的:探讨四周尾部悬吊模拟失重大鼠颈总动脉基质金属蛋白酶(matrix metalloprotein9,MMP9)和金属蛋白酶组织抑制剂1(tissue inhibitor of metalloproteinase1,TIMP1)的基因、蛋白表达及酶活性变化。方法:采用4周(week,wk)尾部悬吊大鼠模拟失重影响,通过透射电镜检测颈总动脉壁基质含量,实时定量聚合酶链式反应检测MMP9和TIMP1的mRNA表达,Western blot和免疫组织化学染色检测其蛋白表达和分布,明胶酶谱法测定MMP9活性水平。结果:与对照组相比,悬吊组大鼠细胞外基质面积较对照组显著增加(P0.05),胶原蛋白含量显著增加(P0.05);悬吊组大鼠颈总动脉MMP9的m RNA表达量无明显改变,而其蛋白表达量和酶活性均显著降低(P0.05);TIMP1 mRNA和蛋白表达量则显著升高(P0.05)。结论:模拟失重使大鼠颈总动脉MMP9水平降低,TIMP1水平升高,可能与其管壁基质增生和胶原蛋白含量增加有关。     

Tissue inhibitor of metalloproteinase‐2 (TIMP‐2) regulates neuromuscular junction development via a β1 integrin‐mediated mechanism

Extracellular matrix (ECM) molecules play critical roles in muscle function by participating in neuromuscular junction (NMJ) development and the establishment of stable, cytoskeleton‐associated adhesions required for muscle contraction. Matrix metalloproteinases (MMPs) are neutral endopeptidases that degrade all ECM components. While the role of MMPs and their inhibitors, the tissue inhibitor of metalloproteinases (TIMPs), has been investigated in many tissues, little is known about their role in muscle development and mature function. TIMP‐2 ^?/? mice display signs of muscle weakness. Here, we report that TIMP‐2 is expressed at the NMJ and its expression is greater in fast‐twitch (extensor digitorum longus, EDL) than slow‐twitch (soleus) muscle. EDL muscle mass is reduced in TIMP‐2^?/? mice without a concomitant change in fiber diameter or number. The TIMP‐2^?/? phenotype is not likely due to increased ECM proteolysis because net MMP activity is actually reduced in TIMP‐2^?/? muscle. Most strikingly, TIMP‐2 colocalizes with β1 integrin at costameres in the wild‐type EDL and β1 integrin expression is significantly reduced in TIMP‐2^?/? EDL. We propose that reduced β1 integrin in fast‐twitch muscle may be associated with destabilized ECM‐cytoskeletal interactions required for muscle contraction in TIMP‐2^?/? muscle; thus, explaining the muscle weakness. Given that fast‐twitch fibers are lost in muscular dystrophies and age‐related sarcopenia, if TIMP‐2 regulates mechanotransduction in an MMP‐independent manner it opens new potential therapeutic avenues. © 2006 Wiley Periodicals, Inc. J Neurobiol, 2006     

Serum tissue inhibitor of metalloproteinase‐1 and risk of cognitive impairment after acute ischaemic stroke

The expression of tissue inhibitor metalloproteinase‐1 (TIMP‐1) significantly increased after acute cerebral ischaemia and involved in neurodegeneration. The purpose was to prospectively investigate the relationship between serum TIMP‐1 with post‐stroke cognitive impairment. Our participants were from an ancillary study of China Antihypertensive Trial in Acute Ischemic Stroke. 598 ischaemic stroke patients from seven participating hospitals were included. Cognitive impairment was evaluated using Mini‐Mental State Examination (MMSE) and Montreal Cognitive Assessment (MoCA) at 3 months. 316 (52.84%) or 384 (64.21%) participants had cognitive impairment according to MMSE or MoCA, respectively. Compared with the first quartile of TIMP‐1, the multivariate‐adjusted odds ratios (95% confidence intervals) for the highest quartile were 1.80 (1.09‐2.97) for cognitive impairment defined by MMSE and 2.55 (1.49‐4.35) by MoCA. Multiple‐adjusted spline regression models showed linear associations between TIMP‐1 concentrations and cognitive impairment (P value for linearity < 0.01). The addition of TIMP‐1 to models including conventional factors improved reclassification for cognitive impairment, as shown by net reclassification index or integrated discrimination improvement (P < 0.05). Participants with both higher TIMP‐1 and matrix metalloproteinase‐9 levels simultaneously had highest risk of cognitive impairment. Higher serum TIMP‐1 levels were associated with increased risk of cognitive impairment after acute ischaemic stroke, independently of established risk factors.     

Neuroprotection conferred by post‐ischemia ethanol therapy in experimental stroke: an inhibitory effect on hyperglycolysis and NADPH oxidase activation

Ethanol provides neuroprotection following ischemia/reperfusion. This study assessed ethanol's effect on hyperglycolysis and NADPH oxidase (NOX) activation. Adult, male Sprague–Dawley rats were subjected to middle cerebral artery occlusion (MCAO) for 2 h. Three sets of experiments were conducted to determine ethanol's effect on (i) conferring neuroprotection by measuring infarct volume and neurological deficits 24 h post reperfusion; (ii) cerebral glucose metabolism and lactic acidosis by measuring brain and blood glucose concentrations and protein expression of glucose transporter 1 and 3 (GLUT1, GLUT3), phosphofructokinase (PFK), as well as lactic acidosis by measuring lactate dehydrogenase (LDH), and lactate; and (iii) nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) activation by detecting enzymatic activity and subunit expression at 3 h after reperfusion. When administered upon reperfusion, ethanol (1.5 g/kg) reduced infarct volume by 40% (p < 0.01) and neurological deficits by 48% at 24 h post reperfusion while reducing (p < 0.01) elevations in glycolytic protein expression and lactate levels during early reperfusion (3 h). Ethanol increased the reductions in cerebral glucose concentration at 3 h post reperfusion by 64% (p < 0.01) while enhancing (p < 0.01) post stroke blood glucose concentration, suggesting a reduced cellular glucose uptake and utilization. Ethanol decreased (p < 0.01) stroke‐induced NOX activation by reducing enzymatic activity and gp91^phox expression by 45% and 38%, respectively. Post‐ischemia ethanol treatment exerts neuroprotection through attenuation of hyperglycolysis and associated NOX activation. Because of the lack of associated hypoglycemia and selectivity toward decreasing cerebral metabolism, further investigation of ethanol's use as a post‐stroke therapy, especially in the context of hyperglycemia, seems warranted.     

PCSK9 inhibitor and atorvastatin reduce cardiac impairment in ovariectomized prediabetic rats via improved mitochondrial function and Ca2+ regulation

Post‐menopausal women have a higher risk of developing cardiometabolic dysfunction. Atorvastatin attenuates dyslipidaemia and cardiac dysfunction but it can have undesirable effects including increased risk of diabetes and myalgia. Currently, the proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitor efficiently reduces low‐density lipoprotein cholesterol (LDL‐C) levels more effectively than atorvastatin. We have been suggested that PCSK9 inhibitor attenuated cardiometabolic impairment more effectively than atorvastatin in ovariectomized prediabetic rats. Female Wistar rats (n = 48) were fed a normal diet (ND) or high‐fat diet (HFD) for 12 weeks. Then, HFD rats were assigned to a sham‐operated (Sham) or ovariectomized (OVX) group. Six weeks after surgery, the OVX group was subdivided into 4 treatment groups: vehicle (HFOV), atorvastatin (HFOA) (40 mg/kg/day; s.c.), PCSK9 inhibitor (HFOP) (4 mg/kg/day; s.c.) and oestrogen (HFOE₂) (50 µg/kg/day; s.c.) for an additional 3 weeks. Metabolic parameters, cardiac and mitochondrial function, and [Ca²⁺]_i transients were evaluated. All HFD rats became obese‐insulin resistant. HFS rats had significantly impaired left ventricular (LV) function, cardiac mitochondrial function and [Ca²⁺]_i transient dysregulation. Oestrogen deprivation (HFOV) aggravated all of these impairments. Our findings indicated that the atorvastatin, PCSK9 inhibitor and oestrogen shared similar efficacy in the attenuation in cardiometabolic impairment in ovariectomized prediabetic rats.     

Extracellular matrix production and regulation in micropatterned endothelial cells

Production and maintenance of extracellular matrix (ECM) is an essential aspect of endothelial cell (EC) function. ECM surfaces composed of collagen type IV and laminin support an atheroprotective endothelium, while fibronectin may encourage an atheroprone endothelium through inflammation or wound repair signaling. ECs maintain this underlying structure through regulation of protein production and degradation, yet the role of cytoskeletal alignment on this regulation is unknown. To examine the regulation and production of ECM by ECs with an atheroprotective phenotype, ECs were micropatterned onto lanes, which created an elongated EC morphology similar to that seen with unidirectional fluid shear stress application. Collagen IV and fibronectin protein production were measured as were gene expression of collagen IV, fibronectin, laminin, MMP2, MMP9, TIMP1, TIMP2, and TGF-β1. ECs were also treated with TNF to simulate an injury model. Micropattern-induced elongation led to significant increases in collagen IV and fibronectin protein production, and collagen IV, laminin, and TGF-β1 gene expression, but no significant changes in the MMP or TIMP genes. TNF treatment significantly increased collagen IV gene and protein production. These results suggest that the increase in ECM synthesis in micropattern-elongated ECs is likely regulated with TGF-β1, and this increase in ECM could be relevant to the atheroprotection needed for maintenance of a healthy endothelium in vivo.     

Oral microcirculation in post-menopause: a possible correlation with periodontitis

doi: 10.1111/j.1741‐2358.2011.00608.x Oral microcirculation in post‐menopause: a possible correlation with periodontitis Objectives: The reduction in the level of oestrogen, typical in menopause, has some effect on the health of the oral cavity. In fact, post‐menopausal women present more severe periodontal disease than pre‐menopausal women. Numerous factors can be held to be responsible for this increase, among which are the effects of oestrogens on the oral epithelium, on the salivary glands, on bone tissue and on the endothelium. Our double blind study aims to evaluate the possible variations in oral microcirculation in post‐menopausal women. Methods: Twenty‐seven women in post‐menopause (age: Mean ± SD: 57.3 ± 8.73) and 27 women in pre‐menopause (age: Mean ± SD: 27.77 ± 3.56) were examined. Oral microcirculation was investigated using oral videocapillaroscopy. Results: The study showed significant differences between cases and controls for the following parameters: decrease in diameter of loops (mean ± SD: 0.038 ± 0.008; 0.045 ± 0.005), increase in tortuosity (mean ± SD: 3.83 ± 1.13; 1.83 ± 1.06) in labial mucosa and decrease in density in periodontal mucosa (Mean ± SD: 28.86 ± 10.92; 89.62 ± 17.83). Conclusion: The decrease in periodontal density may compromise the epithelium tropism, making it prone to inflammation. The tortuosity may indicate a greater permanence of inflammatory factors, increased in post‐menopausal women.     

Peripheral blood flow in menopausal women who have hot flushes and in those who do not.

OBJECTIVE--To compare blood pressure, heart rate, and peripheral vascular responsiveness in menopausal women who have hot flushes and in those who do not, and to assess the effect on these variables of treating women who have hot flushes with oestriol, a natural oestrogen, given vaginally. DESIGN--An open, non-randomised cohort study of flushing and non-flushing menopausal women. A before and after investigation of the effects of vaginal oestriol treatment on the circulation. SETTING--Referral based endocrinology clinic. PATIENTS--88 Consecutive menopausal women, 63 complaining of frequent hot flushes and 25 who had not flushed for at least a year. INTERVENTION--Treatment with vaginal oestriol 0.5 mg at night for six weeks in 18 of the women who had hot flushes. MEASUREMENTS AND MAIN RESULTS--Peripheral blood flow was measured by venous occlusion plethysmography at rest and in response to stressful mental arithmetic and anoxic forearm exercises. Blood flow in the forearm and its variability were significantly higher in flushing than in non-flushing women (4.1 (SD 1.7) and 3.1 (0.9) ml/100 ml tissue/min and 17% and 13% respectively). Blood pressure, heart rate, and blood flow in the hand were, however, similar in the two groups. No difference was found in the peripheral incremental response to either stress or anoxic exercise. Vaginal oestriol significantly lowered forearm blood flow from 4.4 (1.5) to 3.3 (1.1) ml/100 ml tissue/min but dilator responsiveness was unaffected. CONCLUSIONS--The peripheral circulation is different in menopausal women who have hot flushes compared with those who do not, with selective vasodilatation in the forearm. The lowered blood flow in the forearm after vaginal oestriol in flushing women may be relevant to the alleviation of vasomotor symptoms induced by oestrogen treatment.     

MMP‐2 and MMP‐9 as prognostic markers for the early detection of urinary bladder cancer

The present study assessed protein and gene expression levels of tissue inhibitor of metalloproteinase‐2 (TIMP‐2), matrix metalloproteinase‐2 (MMP‐2), and MMP‐9 in urine and blood samples of 50 patients with bladder carcinoma. The expression of TIMP‐2, MMP‐2, and MMP‐9 levels with tumor stage and grade was also assessed. Results showed that the expression levels of MMP‐2 and MMP‐9 in both blood and urine were significantly elevated in group 1 when compared with groups 2 and 3 healthy subjects. The discriminatory ability in the diagnosis of bladder carcinoma of MMP‐2 and MMP‐9 expression was confirmed by receiver operating characteristic curve analysis that revealed a sensitivity and specificity of 100%. MMP‐2 and MMP‐9 levels were not correlated with grade or stage of the tumor. With respect to TIMP‐2 blood and urine levels, results showed a significant decrease in gene expression levels in bladder carcinoma group, whereas, TIMP‐2 protein showed a significant increase in bladder carcinoma.     

Targeted migration of pherophorin‐S indicates extensive extracellular matrix dynamics in Volvox carteri

Hydroxyproline‐rich glycoproteins (HRGPs) constitute a major group of proteins of the extracellular matrix (ECM). The multicellular green alga Volvox carteri is a suitable model organism in which to study the evolutionary transition to multicellularity, including the basic principles and characteristics of an ECM. In Volvox, the ECM is dominated by a single HRGP family: the pherophorins. Our inventory amounts to 117 pherophorin‐related genes in V. carteri. We focused on a pherophorin with an unexpected characteristic: pherophorin‐S is a soluble, non‐cross‐linked ECM protein. Using transformants expressing a YFP‐tagged pherophorin‐S we observed the synthesis and secretion of pherophorin‐S by somatic cells in vivo, and we then traced the protein during its conspicuous migration to the ECM around prehatching juveniles and its localized concentration there. Our results provide insights into how an ECM zone surrounding the progeny is remotely affected by distantly located parental somatic cells. In view of the properties and migration of pherophorin‐S, we conclude that pherophorin‐S is likely to act as an ECM plasticizer to allow for dynamic ECM remodeling.     

Nitric oxide‐mediated regulation of ‐amyloid clearance via alterations of MMP‐9/TIMP‐1

Fibrillar amyloid plaques are largely composed of amyloid‐beta (Aβ) peptides that are metabolized into products, including Aβ1‐16, by proteases including matrix metalloproteinase 9 (MMP‐9). The balance between production and degradation of Aβ proteins is critical to amyloid accumulation and resulting disease. Regulation of MMP‐9 and its endogenous inhibitor tissue inhibitor of metalloproteinase (TIMP)‐1 by nitric oxide (NO) has been shown. We hypothesize that nitric oxide synthase (NOS2) protects against Alzheimer's disease pathology by increasing amyloid clearance through NO regulation of MMP‐9/TIMP‐1 balance. We show NO‐mediated increased MMP‐9/TIMP‐1 ratios enhanced the degradation of fibrillar Aβ in vitro, which was abolished when silenced for MMP‐9 protein translation. The in vivo relationship between MMP‐9, NO and Aβ degradation was examined by comparing an Alzheimer's disease mouse model that expresses NOS2 with a model lacking NOS2. To quantitate MMP‐9 mediated changes, we generated an antibody recognizing the Aβ1‐16 fragment, and used mass spectrometry multi‐reaction monitoring assay for detection of immunoprecipitated Aβ1‐16 peptides. Aβ1‐16 levels decreased in brain lysates lacking NOS2 when compared with strains that express human amyloid precursor protein on the NOS2 background. TIMP‐1 increased in the APPSwDI/NOS2^?/? mice with decreased MMP activity and increased amyloid burden, thereby supporting roles for NO in the regulation of MMP/TIMP balance and plaque clearance.