Cytoplasm Of Smooth Muscle Cells Research Articles

Low androgen status inhibits erectile function by increasing pyroptosis in rat corpus cavernosum

The mechanism of erectile dysfunction (ED) caused by low androgen status is not fully understood. To investigate whether low androgen status inhibits erectile function of rats by inducing pyroptosis in the corpus cavernosum (CC). Thirty-six eight-weeks-old healthy male Sprague-Dawley rats were equally divided into six groups: sham-operated group (4w sham, 8w sham), castration group (4w cast, 8w cast), and castration + testosterone (T) group (4w cast + T, 8w cast + T). The rats in castration + T groups were injected with testosterone propionate subcutaneously every other day. After 4 and 8weeks, the ratio of maximum intracavernous pressure (ICPmax)/mean arterial pressure (MAP), the level of serum T, the concentration of nitric oxide (NO) and interleukin-1β (IL-1β), the expression of NOD-like receptor pyrin domain containing 3 (NLRP3), apoptosis-associated speck-like protein containing a caspase activation and recruitment domain (ASC), Caspase-1 p20, gasdermin D-N (GSDMD-N), transforming growth factor β1 (TGF-β1), collagen-I, and collagen-III, the ratio of smooth muscle/collagen (SM/C), and the proportion of pyroptotic cells in the CC were analyzed. The ratio of ICPmax/MAP (3/5V) and SM/C, the level of NO and serum T was significantly decreased in castration groups when compared to other groups (p<0.01). NLRP3, ASC, Caspase-1, and GSDMD were mainly expressed in the cytoplasm of smooth muscle cells (SMCs) and endothelial cells (ECs) in the CC. The expression of NLRP3, ASC, Caspase-1p20, GSDMD-N, IL-1β, TGF-β1, collagen-I, and collagen-III was significantly increased in castration groups when compared with other groups (p<0.01). The proportion of pyroptotic cells in the CC was increased significantly in castration groups when compared with other groups (p<0.05). Low androgen status inhibits erectile function of rats by promoting CC fibrosis and reducing NO synthesis through pyroptosis of SMCs and ECs in the CC.

Brown Bowel Syndrome: A Multi-institutional Case Series.

Brown bowel syndrome (BBS) is a rare condition associated with vitamin E deficiency and defined by prominent lipofuscin deposition in the muscularis propria. Eight unique cases of BBS were identified: 5 men and 3 women (mean age=58.6 y). Pertinent comorbidities included bariatric surgery=2, malnourishment=2, Crohn=2, cystic fibrosis=1, alcohol and cocaine abuse=1, and prior small bowel resections=1. Presenting symptoms included abdominal pain=3, bleeding=1, nausea and vomiting=1, and nonresponsiveness=1. Imaging studies were often abnormal: thickened bowel wall=3 (1 with a mass), small bowel obstruction=2, and edematous and dilated bowel wall=2. Most specimens were surgical resections (n=7, autopsy=1): extended right colectomy=2, small bowel only=5 (terminal ileum=3, jejunum=2). Two specimens were grossly described as mahogany, and 1 case contained a perforation. Histologic sections of all cases showed finely granular, brown cytoplasmic pigment in smooth muscle cells on hematoxylin and eosin. This pigment was most conspicuous in the muscularis propria (small bowel>colon), and it was not identified in the mucosa. The pigment was reactive with Fontana-Masson, carbol lipofuscin, Periodic acid-Schiff, and Periodic acid-Schiff with diastase, and electron microscopy was compatible with lipofuscin. The mean clinical follow-up was 208 weeks: 1 patient died of complications of encephalitis, the others were alive and well. BBS is important to recognize because it is linked with malnutrition, specifically vitamin E deficiency, and it can (rarely) clinically simulate malignancy. The diagnosis is based on the identification of the lipofuscin pigment in the cytoplasm of smooth muscle cells, which is most easily seen in the muscularis propria of the small bowel.

Polyamine concentration is increased in thoracic ascending aorta of patients with bicuspid aortic valve.

Polyamines are cationic molecules synthesized via a highly regulated pathway, obtained from the diet or produced by the gut microbiota. They are involved in general molecular and cellular phenomena that play a role also in vascular disease. Bicuspid aortic valve (BAV) is a congenital malformation associated to a greater risk of thoracic ascending aorta (TAA) aneurysm, whose pathogenesis is not yet well understood. We focused on differential analysis of key members of polyamine pathway and on polyamine concentration in non-dilated TAA samples from patients with either stenotic tricuspid aortic valve (TAV) or BAV (diameter≤45mm), vs. normal aortas from organ donors, with the aim of revealing a potential involvement of polyamines in early aortopathy. Changes of gene expression in TAA samples were evaluated by RT-PCR. Changes of ornithine decarboxylase 1 (ODC1), a key enzyme in polyamine formation, and cationic amino acid transporter 1 (SLC7A1/CAT-1) expression were analyzed also by Western blot. ODC1 subcellular localization was assessed by immunohistochemistry. Polyamine concentration in TAA samples was evaluated by HPLC. BAV TAA samples showed an increased concentration of putrescine and spermidine vs. TAV and donor samples, together with a decreased mRNA level of polyamine anabolic enzymes and of the putative polyamine transporter SLC7A1/CAT-1. The catabolic enzyme spermidine/spermine N1-acetyltransferase 1 showed a significant mRNA increase in TAV samples only, together with a decreased concentration of spermine. The decreased expression of SLC7A1/CAT-1 and ODC1 mRNAs in BAV corresponded to increased or unchanged expression of the respective proteins. ODC was located mainly in smooth muscle cell (SMC) nucleus in TAV and donor samples, while it was present also in SMC cytoplasm in BAV samples, suggesting its activation. In conclusion, BAV, but not TAV non-dilated samples show increased polyamine concentration, accompanied by the activation of a regulatory negative feedback mechanism.

The role of MIF, cyclinD1 and ERK in the development of pulmonary hypertension in broilers

ABSTRACTPulmonary hypertension (PH) is a major disease in the broiler breeding industry. During PH, the pulmonary artery undergoes remodelling, which is caused by pulmonary vascular smooth muscle cell proliferation. CyclinD1 regulates cell proliferation. This study investigated the role of cyclinD1 in the development of PH in broilers, and which bioactivators and signalling pathway are involved in the pathological process. The PH group contained 3–4-week-old broilers with clinical PH, and the healthy group broilers from the same flock without PH. Histopathology indicated pulmonary arterial walls were thicker in the PH group compared with the healthy group. Target gene expressions of macrophage migration inhibitory factor (MIF), extracellular signal-regulated kinase (ERK), and cyclinD1 detected by quantitative real-time PCR were upregulated in the PH group compared with the healthy group. Immunohistochemistry showed MIF, phosphorylated ERK (p-ERK) and cyclinD1 were present on pulmonary vascular walls; MIF was present in the cytoplasm of arterial endothelial cells and smooth muscle cells; p-ERK and cyclinD1 were present in smooth muscle cell cytoplasm. Western blotting demonstrated that MIF, p-ERKand cyclinD1 levels were significantly higher (P < 0.01) in the PH group compared with the healthy group. In summary, increased MIF in PH broiler pulmonary arteries upregulated cyclinD1 via the ERK signalling pathway to induce pulmonary vascular smooth muscle cell proliferation, causing pulmonary artery remodelling and hypertension.

Morphological changes in the colonic muscular layer and interstitial cells of Cajal in experimental acute ulcerative colitis

to investigate morphological changes in the muscular layer and an intermuscular population of interstitial cells of Cajal (ICC) in acute ulcerative colitis induced by dextran sulfate sodium (DSS). Acute colitis was induced by 3% DSS (molecular weight 40 kDa) in drinking water for 5 days; a control animal group was given tap water. Longitudinal colonic sections were histologically examined. Immunofluorescence staining of total preparations with antibodies to c-Kit was employed to assess an ICC population. Morphological examination showed a morphological pattern of acute ulcerative colitis in the DSS-treated mice. The colonic muscular layer was thickened and the cytoplasm of smooth muscle cells was vacuolated. There was a decrease in ICC network density and counts in the muscular layer, as well as a specific c-Kit positive granularity in the ICC bodies and processes. In acute ulcerative colitis, there are alternative changes in the colonic muscular layer and a decrease in ICC density.

Expression and significance of NELIN and SM22α in varicose vein tissue.

The aim of the present study was to investigate the expression of NELIN and SM22α in lower extremity varicose vein tissue, and their association with varicose veins. Tissue samples were collected from 18 patients with lower extremity varicose veins for the experimental group, while normal great saphenous vein tissue was reserved during coronary artery bypass surgery from 14 patients for the controls. Reverse transcription polymerase chain reaction (RT-PCR) analysis was applied to detect the mRNA expression levels of NELIN and SM22α, while immunohistochemical techniques were used to detect the protein expression levels in the normal and abnormal veins. RT-PCR results revealed that the mRNA expression levels of NELIN and SM22α in the experimental group decreased significantly when compared with the control group (P<0.01). In the two groups, immunohistochemical staining demonstrated that NELIN and SM22α were primarily expressed in the cytoplasm of smooth muscle cells, and the expression quantity decreased significantly in the experimental group when compared with the control group (P<0.05). The low expression of SM22α in the primary lower limb varicose vein tissue indicated that the vascular smooth muscle cell layer had transformed from a contractile to a secretory phenotype, which may have resulted in the remodeling of the vein walls and the occurrence of varicose veins. Therefore, NELIN and SM22α were demonstrated to play a key role in the development of varicosity.

Calmodulin inhibitors suppress calcium signaling from serotonin receptors in smooth muscle cells and abolish vasoconstrictive response on intravenous introduction of serotonin

Comparative study of the effect of calmodulin inhibitors (trifluoperazine, W-12, and W-13) and the TRPVI channel blocker (capsazepine) on receptor-dependent calcium exchange in smooth muscle cells of the rat aorta and on the contractility of the isolated aorta was conducted. It was determined that trifluoperazine almost completely removes an increase in the concentration of calcium ions in the cytoplasm of smooth muscle cells (isolated from the rat aorta) and smooth muscle cells of the A7r5 line in response to serotonin and does not influence the cell response to vasopressin and angiotensin II. W-12 and W-13 also do not reduce calcium ion concentration increase (induced by vasopressin and angiotensin II) but reduces by two times its rise in response to serotonin. It was found that the efficiency of calcium exchange suppression by calmodulin inhibitors correlates with the intensity at which they inhibit the contractile response of the aorta on the effect of serotonin. It was detected that the inhibiting effect of calmodulin blockers on calcium exchange in smooth muscle cells and the contractility of the rat isolated aorta during the activation of serotonin vasoconstrictive receptors are realized by a TRPV1-independent mechanism. It was demonstrated in experiments in vivo that trifluoperazine does not influence hypotensive reaction in rats (normally observed in response to intravenous serotonin introduction), but removes the hypertensive effect of this neurotransmitter in rats after chronic introduction of dexamethasone. The results obtained confirm the hypothesis (that we previously stated) about the direct involvement of calmodulin in signal transmission from vasoconstrictive serotonin receptors.

VIP/PACAP receptors in cerebral arteries of rat: Characterization, localization and relation to intracellular calcium

BackgroundVasoactive intestinal peptide (VIP) and pituitary adenylate cyclase activating peptide (PACAP)-containing nerves surround cerebral blood vessels. The peptides have potent vasodilator properties via smooth muscle cell receptors and activation of adenylate cyclase. The purpose of this study was to describe the effects of two putative VIP/PACAP receptor antagonists and the distribution of the receptor protein in rat brain vessels. MethodsThe vascular effects of VIP, PACAP-27 and PACAP-38 were investigated in segments of rat middle cerebral artery (MCA) by pressurized arteriography, and in a wire myograph. The antagonistic responses to PACAP6-38 and PG99-465 were evaluated. In addition, the receptor subtypes for VIP and PACAP (VPAC1, VPAC2 and PAC1) were visualized in the rat middle cerebral artery by immunohistochemistry and Western blotting. ResultsIn the perfusion model, abluminal but not luminal VIP, PACAP-27 and PACAP-38 caused concentration-dependent relaxations of the MCA (27.1±0.2%, 25.2±0.4% and 0.3±0.1%, respectively). In the wire myograph, there was no significant difference in potency of the peptides in the MCA. In both systems, PACAP6-38 and PG99-465 inhibited the VIP induced relaxation. Western blot showed the presence of the receptor proteins in cerebral vasculature and immunohistochemistry showed that all three receptors are present and located in the cytoplasm of smooth muscle cells. ConclusionIn both systems, the two blockers antagonized the relaxant VIP effect; the potency order of agonists and the immunohistochemistry suggest the presence of the dilatory VPAC1 and VPAC2 receptors on the smooth muscle cells.

C-Type Natriuretic Peptide Specifically Acts on the Pylorus and Large Intestine in Mouse Gastrointestinal Tract

C-type natriuretic peptide (CNP) exerts its main biological effects by binding to natriuretic peptide receptor B (NPR-B), a membrane-bound guanylyl cyclase receptor that produces cyclic guanosine monophosphate (cGMP). CNP is known to cause gastrointestinal (GI) smooth muscle relaxation. Experimental evidence suggests a connection between CNP signaling and GI function, with reactive regions in the GI tract possibly affecting transit; however, this relation has not yet been conclusively shown. Here, we show that CNP plays important region-specific roles in the GI tract of mice. We found that treatment with CNP (1 or 2 mg/kg) increased transient cGMP production in the pylorus, colon, and rectum, with the higher dose (2 mg/kg) enhancing gastric emptying in mice; this increase in cGMP levels was however absent in NPR-B-deficient short-limbed dwarfism (SLW) mouse. Furthermore, we found that NPR-B is highly expressed in the pylorus, colon, and rectum, being localized to nerve fibers and to the nuclei and cytoplasm of smooth muscle cells of the GI tract and blood vessels. Our invivo findings showed that NPR-B-mediated cGMP production after CNP administration specifically acted on the pylorus, colon, and rectum and contributed to gastric emptying. CNP may thus be a potential therapeutic agent for GI motility/transit disorders such as ileus and pyloric stenosis.

Expression of smoothelin and smooth muscle actin in the skin.

Smoothelin is a cytoskeletal protein of differentiated smooth muscle cells with contractile capacity, distinguishing it from other smooth muscle proteins, such as smooth muscle actin (SMA). To evaluate the expression of smoothelin and SMA in the skin in order to establish specific localizations of smoothelin in smooth muscle cells with high contractile capacity and in the epithelial component of cutaneous adnexal structures. Immunohistochemical analysis (smoothelin and SMA) was performed in 18 patients with normal skin. SMA was expressed by the vascular structures of superficial, deep, intermediate and adventitial plexuses, whereas smoothelin was specifically expressed in the cytoplasm of smooth muscle cells of the deepest vascular plexus and in no other plexus of the dermis. The hair erector muscle showed intense expression of smoothelin and SMA. Cells with nuclear expression of smoothelin and cytoplasmic expression of SMA were observed in the outer root sheath of the inferior portion of the hair follicles and intense cytoplasmic expression in cells of the dermal sheath to SMA. We report the first study of smoothelin expression in normal skin, which differentiates the superficial vascular plexus from the deep. The deep plexus comprises vessels with high contractile capacity, which is important for understanding dermal hemodynamics in normal skin and pathological processes. We suggest that the function of smoothelin in the outer root sheath may be to enhance the function of SMA, which has been related to mechanical stress. Smoothelin has not been studied in cutaneous pathology; however we believe it may be a marker specific for the diagnosis of leiomyomas and leiomyosarcomas of the skin. Also, smoothelin could differentiate arteriovenous malformations of cavernous hemangioma of the skin.

Downregulation of FHL1 Expression in Thoracic Aortic Dissection: Implications in Aortic Wall Remodeling and Pathogenesis of Thoracic Aortic Dissection

To investigate the potential role of four and a half Lin-11, Isl-1, and Mec-3 (FHL1) protein in the pathogenesis of thoracic aortic dissection (TAD). The expression levels and localization of FHL1 protein in aortic tissue of TAD were analyzed using Western blot and immunohistochemistry. Furthermore, small interfering ribonucleic acid was used to knock down the FHL1 gene in rat aortic smooth muscle cells (SMCs). After assessing knockdown efficiency and specificity by real-time polymerase chain reaction and Westernblot, the effect of FHL1 knockdown on cell proliferation and apoptosis was evaluated by 3- (4,5-Dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide assay and flow cytometry, respectively. Compared with normal aortic tissue, FHL1 protein expression in aortic tissue from TAD patients was significantly downregulated. Immunohistochemistry analysis showed that FHL1 was mainly localized in the cytoplasm of SMCs. In diseased aortic tissue, FHL1 immunoreactivity was lowest in SMCs in the split aortic media and adjacent area, but relatively high in SMCs in the aortic intima and adventitia. FHL1 knockdown significantly inhibited the proliferation of rat aortic SMCs but exerted no obvious effect on cell apoptosis. FHL1 protein expression is downregulated in TAD. Downregulation of FHL1 expression might contribute to the pathogenesis of TAD, perhaps by suppressing the proliferation of aortic SMCs and affecting aortic wall remodeling.

Regulation of the lumen of a resistance blood vessel by mechanical stimuli

The flow in a vessel able to regulate its lumen under the action of mechanical stimuli, the variation of the pressure difference between the inner and outer surfaces of the vessel wall and the blood flow velocity, is described. This ability is determined by the effect of the mechanical stimuli on the degree of activation of smooth muscle cells in the vessel wall. In order to describe the active properties of the wall, two controlling parameters which have the sense of the concentration of free calcium ions in the cytoplasm of smooth muscle cells and the average concentration of nitric oxide in the smooth muscle layer, are introduced. The approach proposed makes it possible to estimate both the degree of participation of each mechanical stimulus in vessel lumen regulation and the result of interaction of two differently directed vascular responses. The calculations show that both the magnitude and direction of the radius response to a mechanical stimulus depend on the initial state of the vessel wall. The role of the vessel wall sensitivity to mechanical stimuli in the stabilization of the blood flow-rate and the variation of the radius along the vessel is considered.

Characterization of VEGF and Angiopoietins Expression in Human Corpus Cavernosum during Aging

Erectile dysfunction (ED) is a highly prevalent and age-related disease, caused by endothelial dysfunction and impaired cavernous angiogenesis. However, cellular and molecular changes involved in erectile pathophysiology in aging male remain to be elucidated. To characterize the vascular organization, concomitantly with analysis of the expression of vascular endothelial growth factor (VEGF), Angiopoietin 1 (Ang1) and Angiopoietin 2 (Ang2) in young and aged human corpus cavernosum. Human penile fragments were removed from patients submitted to penile deviation surgery (11 cases; 58-70 years) and from potential organ donors (four cases; 18-28 years) without ED or risk factors for ED. Smooth muscle and connective tissue were assessed by Masson's trichrome staining and computer-assisted histomorphometry. Dual immunostaining for specific markers of endothelium (von Willebrand factor) and smooth muscle cell (alpha-actin), VEGF, Ang1 and Ang2 was assayed by fluorescence microscopy. Semi-quantification of expression of angiogenic factors was performed by Western blotting. Expression of VEGF and Angiopoietins in human corpus cavernosum, using a combination of histologic stainings, and molecular biology tools in order to achieve a better understanding of cavernosal tissue remodeling with aging. Aged human corpus cavernosum presented wider sinusoidal spaces, loss of muscle cell bundles, and increased connective tissue content. Ang1 was scarcely expressed in small clusters in smooth muscle cell cytoplasm with identical localization in both studied groups. VEGF expression was abundant in smooth muscle cell and its expression markedly decreased in aged tissue, contrasting with the expression of angiopoietins that increased in the aged corpus cavernosum. Immunoflourescent studies of cellular markers and growth factors help clarifying vascular organization and angiogenesis mechanisms in erectile tissue. Our findings demonstrate that the organization pattern of vascular endothelium and smooth muscle components of cavernosal tissue modifies during aging. Ang1 and Ang2 upregulation in human-aged penile tissue suggest a VEGF-independent vascular remodeling mechanism.

Preparation,identification and application of polyclonal antibodies against human PIWIL2 protein

Objective:To prepare rabbit polyclonal antibody against human P-element-induced wimpy testis like 2 (HIWI2) protein,identify its properties and investigate its distribution in normal and tumor tissues by means of tissue chip. Methods: PIWIL2 peptide was synthesized chemically and conjugated to Keyhole limpet hemocyanin (KLH) as an immunogen. Then the PIWIL2-KLH conjugation was injected into rabbits subcutaneously to produce polyclonal antibodies. The specificity and sensitivity of antibodies were identified by ELISA and Western blotting after purification by affinity chromatography. PIWIL2 was then immuno-stained on the tissue chip to study its distribution in the normal and tumor tissues. Results: Rabbit’s antibodies against human PIWIL2 were prepared after the injection of PIWIL2-KLH conjugation subcutaneously. These antibodies were identified as PIWIL2 peptides by ELISA and Western blotting assay. PIWIL2 protein expression was tissue-specific in tumor tissues,with PIWIL2 protein found in the cytoplasm of smooth muscle cells of most normal and tumor tissues. Conclusion: The polyclonal antibodies against human PIWIL2 protein have been successfully prepared,which provides a basis for further study on the role of PIWIL2 in the pathway of miRNA/RNA.

Expression of COX-2 and pregnancy associate plasma protein A in coronary arteries and their relationship with acute coronary syndrome: an autopsy study of 42 cases

To study the expression of COX-2 and pregnancy associate plasma protein A (PAPP-A) in coronary arteries and their relationship with acute coronary syndrome. Twenty-one autopsy cases with acute coronary syndrome encountered during the period from 2002 to 2007 were enrolled into the study. Another 21 autopsy cases without evidence of acute coronary syndrome were used as the controls. The right and left coronary arteries of each group were dissected, embedded and processed as paraffin sections. Immunohistochemical study for CD68 and alpha-actin was performed to highlight the presence of macrophages and smooth muscle cells, respectively. The expression of COX-2 and PAPP-A was evaluated. In the acute coronary syndrome group, COX-2 was localized mainly in the cytoplasm of endothelial cells, macrophages and smooth muscle cells. COX-2 expression in the cytoplasm of smooth muscle cells (28.60%) was significantly higher than that in the control group (4.76%, chi(2) = 14.13, P< 0.05). There was a positive correlation on COX-2 and PAPP-A expression in smooth muscle cells of the media layer of coronary arteries in acute coronary syndrome group (r = 0.88, P < 0.05). The expression of PAPP-A in smooth muscle cells of the media layer in coronary arteries not associated with plaque formation, was higher than that when there were atherosclerotic plaques (chi(2) = 10.36, P < 0.05). In coronary arteries, COX-2 and PAPP-A play certain roles in the pathogenesis of acute coronary syndrome.

Masson Trichrome Stain Helps Differentiate Myofibroma from Smooth Muscle Lesions in the Head and Neck Region

Myofibromas are well described in the head and neck region, but differentiating them from smooth muscle lesions is still difficult using smooth muscle immunohistochemical stains. This study evaluated the usefulness of the Masson trichrome stain in differentiating myofibromas from smooth muscle lesions in the head and neck region. Samples of 11 oral myofibromas, two leiomyomas, one angioleiomyoma, and one smooth muscle hamartoma were retrieved from our archives. Immunohistochemistry and Masson trichrome stains were performed on tissue sections of these lesions. All 11 oral myofibromas, seven from male patients and four from female patients, were solitary myofibromas. The patients' mean age at diagnosis was 32.8 years. Oral myofibromas occurred most commonly on the gingiva (four cases) and in the mandible (three cases). With the Masson trichrome stain, the smooth muscle cell cytoplasm was stained red, while the collagenous fibrous tissue was stained blue. Myofibromas and smooth muscle lesions demonstrated different characteristic patterns with the Masson trichrome stain. Myofibromas were composed of a much more collagenous stroma intermixed with the spindle cells. Thick fibrous bundles with random, irregularly intersecting angles were prominent in myofibromas. Smooth muscle lesions showed only minimal delicate fibrous tissue surrounding the smooth muscle cells and in the septa between the smooth muscle masses. On low-power view, red masses of smooth muscle tumor surrounded by blue fibrous tissue were observed. The Masson trichrome stain can be a useful tool to differentiate myofibromas from smooth muscle lesions, but immunohistochemical methods to rule out other spindle cell lesions are still needed.

Gene therapy to promote new blood vessel formation in the ischemic heart: where do we stand?

Erectile dysfunction (ED) is a highly prevalent and age-related disease, caused by endothelial dysfunction and impaired cavernous angiogenesis. However, cellular and molecular changes involved in erectile pathophysiology in aging male remain to be elucidated.To characterize the vascular organization, concomitantly with analysis of the expression of vascular endothelial growth factor (VEGF), Angiopoietin 1 (Ang1) and Angiopoietin 2 (Ang2) in young and aged human corpus cavernosum.Human penile fragments were removed from patients submitted to penile deviation surgery (11 cases; 58–70 years) and from potential organ donors (four cases; 18–28 years) without ED or risk factors for ED. Smooth muscle and connective tissue were assessed by Masson's trichrome staining and computer-assisted histomorphometry. Dual immunostaining for specific markers of endothelium (von Willebrand factor) and smooth muscle cell (alpha-actin), VEGF, Ang1 and Ang2 was assayed by fluorescence microscopy. Semi-quantification of expression of angiogenic factors was performed by Western blotting.Expression of VEGF and Angiopoietins in human corpus cavernosum, using a combination of histologic stainings, and molecular biology tools in order to achieve a better understanding of cavernosal tissue remodeling with aging.Aged human corpus cavernosum presented wider sinusoidal spaces, loss of muscle cell bundles, and increased connective tissue content. Ang1 was scarcely expressed in small clusters in smooth muscle cell cytoplasm with identical localization in both studied groups. VEGF expression was abundant in smooth muscle cell and its expression markedly decreased in aged tissue, contrasting with the expression of angiopoietins that increased in the aged corpus cavernosum.Immunoflourescent studies of cellular markers and growth factors help clarifying vascular organization and angiogenesis mechanisms in erectile tissue. Our findings demonstrate that the organization pattern of vascular endothelium and smooth muscle components of cavernosal tissue modifies during aging. Ang1 and Ang2 upregulation in human-aged penile tissue suggest a VEGF-independent vascular remodeling mechanism. Tomada N, Tomada I, Cruz F, Vendeira P, and Neves D. Characterization of VEGF and angiopoietins expression in human corpus cavernosum during aging.

Association of aryl hydrocarbon receptor and cytochrome P4501B1 expressions in human non-small cell lung cancers

It has been shown that cytochrome P4501B1 (CYP1B1) is over-expressed in variety of tumors. The objective of this study was to examine the expression and localization of CYP1B1 in normal lungs and tumor tissues of non-small cell lung cancers (NSCLC). CYP1B1 was detectable in the microsomes of normal lungs and lung tumors with Western immunoblotting. Utilizing immunohistochemical staining, we found that CYP1B1 was constitutively expressed in the cytoplasm of smooth muscle cells, but not in pneumocytes. Bronchiolar epithelia from three out of 19 normal lungs expressed CYP1B1. Among 89 NSCLC tumor tissues, more than 50% expressed CYP1B1. In addition, CYP1B1 expression was more common in adenocarcinomas than in squamous cell carcinomas. Recently, we reported over-expression of aryl hydrocarbon receptors (AhR) in lung adenocarcinomas. In the present study, we observed an association between CYP1B1 and AhR expression in 89 NSCLC tumor tissues. These results suggest that CYP1B1 overexpression in NSCLC is accompanied by up-regulation of AhR expression.

Redox signaling in oxygen sensing by vessels.

In response to the increase in oxygen tension at birth, the resistance pulmonary arteries dilate, while the ductus arteriosus constricts. Although modulated by the endothelium, these opposite responses are intrinsic to the vascular smooth muscle. While still controversial, it seems likely that during normoxia the production of reactive oxygen species (ROS) increases and the smooth muscle cell cytoplasm is more oxidized in both pulmonary arteries and ductus, compared to hypoxia. However, the effect of changes in the endogenous redox status or the addition of a redox agent, oxidizing or reducing, is exactly opposite in the two vessels. A reducing agent, dithiothreitol, like hypoxia, in the pulmonary artery will inhibit potassium current, cause depolarization, increase cytosolic calcium and lead to contraction. Responses to dithiothreitol in the ductus are opposite and removal of endogenous H(2)O(2) by intracellular catalase in the ductus increases potassium current. Oxygen sensing in both vessels is probably mediated by redox effects on both calcium influx and calcium release from the sarcoplasmic reticulum (SR).

Identification and expression of scavenger receptor SR-BI in endothelial cells and smooth muscle cells of rat aorta in vitro and in vivo

In this study, we used immunoelectron microscopy to investigate the subcellular localization of scavenger receptor class B type I (SR-BI) in the arterial walls of rats. The expression of SR-BI in cultured endothelial and smooth muscle cells of rat aorta after exposure to high-density lipoprotein (HDL) was also investigated by immunofluorescence microscopy and immunoblotting analysis. A peptide containing residues 495–509 from mouse SR-BI (mSR-BI) plus an NH 2-terminal cysteine was coupled to hemocyanin to generate mSR-BI antiserum in rabbits. Reactivity of antiserum against the synthetic peptides was confirmed with an enzyme-linked immunosorbent assay (ELISA). The results showed that SR-BI was specifically localized on the surface of the endothelial cells and smooth muscle cells. SR-BI was also observed in the cytoplasm of smooth muscle cells. Immunoblotting analysis indicated that SR-BI was expressed in the cell membrane. The levels of SR-BI increased gradually from 1 to 3 h and decreased at 24 and 48 h after cholesterol-loaded cells were incubated in the culture medium containing HDL. We conclude that SR-BI, a functional receptor for HDL, is expressed in the aortic endothelial cells as well as in smooth muscle cells. This receptor also responds to the presence of HDL in the culture medium.