Cyclosporine-A (CsA) is the widely used immunosuppressant drug in renal transplantation. However, the effects of cyclosporine-A are limited by a significant nephrotoxicity. The mechanisms of CsA-induced allograft nephropathy are remaining controversial. Recent study indicated that cellular apoptosis may contribute to the cyclosporine A-mediated cytotoxic action. To date, regarding the effects of cyclosporine A on renal cell apoptosis-related gene expression remain poorly defined. p53 is an important gene in control of renal cell growth and death. Transforming growth factor-beta (TGF-beta) is a multifunctional cytokine that has anti-proliferative as well as fibrogenic properties.

We hypothesized that cyclosporine-A may increase p53 and TGF-β expression in renal tubular cells. These actions of cyclosporine-A may contribute to the cellular apoptosis, fibrosis and CsA-induced nephrotoxicity. Therefore, current study was designed to determine the effects of cyclosporine-A on the p53 and TGF-βl protein expression by immunohistochemical staining (IHCS) in cultured human tubular cells.

Esophageal cancer is the fastest growing malignancy of any tumor in the United States with an increased incidence of adenocarcinoma across all socioeconomic boundaries. The stage and the residual tumor classification are the most important prognostic factors. However, patients with similar stage of disease show a marked difference in survival. Therefore, there are needs to identify new molecular prognostic markers that may help to better assess the survival probability.

The recently introduced genetic diagnosis of cancer micrometastasis is quite attractive because of its high detection sensitivity. Not infrequently, however, there are marked discrepancies between genetic and conventional histologic diagnoses, especially concerning lymph nodes from carcinoma patients. Alterations in the p53 tumor suppressor gene are the most frequent genetic changes found in breast and lung cancer. The p53 gene functions as a negative regulator of cell growth. Alterations in the gene lead to loss of its usual negative growth regulation and more rapid cell proliferation.

Endothelin-1 (ET-1) is a potent endothelial cell-drived vasoconstrictive peptide which is increased in congestive heart failure (CHF). ET-1 is converted from its precursor big ET-1 by activation of endothelin converting enzyme (ECE). ET-1 binding to ET-A receptor in vascular smooth muscle cells stimulates vasoconstriction and binding to ET-B receptor in vascular endothelial cells mediates vasodilation. In previous studies, we and others demonstrated that plasma ET-1 was significantly increased in congestive heart failure. However, the presentation and localization of endothelin converting enzyme and endothelin receptors (ET-A and ET-B) in human cardiac tissue with and without heart failure remain unclear. Therefore, the current study was designed to investigate the expression and localization of endothelin receptors and endothelin converting enzyme in human myocardium in the absence or presence of congestive heart failure.

Human atrial tissues (n=6) were obtained from normal subjects and end-stage CHF patients during cardiac transplantation. The expression of ECE, ET-A and ET-B were determined by immunohistochemical staining (IHCS).

Endothelin (ET-1) is a potent renal and systemic vasoconstrictor and sodium regulating peptide. Endothelin synthesis in the kidney have been reported in glomerulus endothelial, epithelial and mesangial cells as well as in inner medullary collecting duct. Factors stimulating the production of endothelin include shear stress, hypoxia, vasoactive agents and cytokines. Endothelin binding to ET-A receptor in vascular smooth muscle cells stimulates vasoconstriction.

Renal graft rejection is a major problem after kidney transplantation with severe renal damage and renal vasoconstriction. We hypothesized that renal tissue level of endothelin-1, endothelin receptors and endothelin converting enzyme (ECE) may increase in renal tissue with rejection after kidney transplantation. Therefore, the current study was designed to determine the endothelin-1 and endothelin receptors (ET-A and ET-B) as well as endothelin converting enzyme level by immunohistochemical staining (IHCS) in human renal tissue with rejection after kidney transplantation.

Transforming growth factor-beta (TGF-β) is a growth-regulating peptide that has been shown to enhance collagen production both in vivo and in vitro. The previous studies demonstrated that TGF-β 1 is present in the normal animal myocardium. However, the expression and localization of TGF-β 1 and TGF-P receptor in human myocardium remain unclear. Therefore, the present study was designed to determine the TGF-β 1 and its receptor in human myocardium in normal subjects and in patients with end-stage congestive heart failure (CHF).

Human ventricular tissues were obtained from five normal subjects and five patients with end-stage CHF during cardiac transplantation. TGF-β 1 and TGF-beta type I receptor (TGF-βRI) were determined by immunohistochemical staining (IHCS). The results of IHCS was evaluated by staining density scores (0, no staining; 1, minimal staining; 2, mild staining; 3, moderate staining; and 4, strong staining). The positive staining area (+%) in entire section was also determined.

Transforming growth factor-bate (TGF-β) is a growth-relating peptide that has been shown to enhance collagen production in vivo and in vitro. TGF-β isoforms include TGF-β 1, β2 and β3. TGF-β receptors subtypes include type I (TGFβRI) and type II (TGFβRII) receptors. Overproduction of TGF-β can lead to real damage.

Renal graft rejection is major problem after kidney transplantation with severe renal damage. We hypothesized that renal tissue level of TGF-β may increase in renal rejection after kidney transplantation. Therefore, the current study was designed to determine the TGF-β 1 isoform and its receptor TGFβRI in human renal tissue with rejection by immunohistochemical staining (IHCS) and compared with normal human renal tissue. The results of IHCS was evaluated by IHCS staining density scores (0, no staining; 1, minimal staining; 2, mild staining; 3, moderate staining; and 4, strong staining). The positive staining area (+%) in entire section was also determined. The sections treated with preabsorbed blocking peptide or nonimmune rabbit serum demonstrated no immunoperoxidase activity.

Nitric oxide (NO) is a potent endothelial-drived vasorelaxing factor and may involved in salt sensitive hypertension and renal damage. Nitric oxide is produced from L-arginine by activation of nitric oxide synthase (NOS). Three NOS has been identified which are brain NOS (bNOS), inducible NOS (iNOS) and endothelial NOS (eNOS). On the other hand, renal graft rejection is major problem after kidney transplantation with severe renal damage and renal vasoconstriction. While NOS has been reported localized in human and animal kidney, the status of three NOS in human renal tissue with rejection remains poorly defined. Therefore, the present study was designed to determine the expression of bNOS, iNOS and eNOS by immunohistochemical staining (IHCS) in human renal tissue with rejection and compared with that in normal human renal tissue.

Human renal biopsy (n=5) were obtained after kidney transplantation with mild and moderate renal rejection. Normal kidney biopsy was obtained during nephrectomy.

Angiotensin II (AH) is a potent vasoconstrictor and mitogenic factor. AH receptors include type 1 (ATI) and type 2 (AT2) receptors. Recent studies demonstrated that both ATI and AT2 receptors expressed in human myocardium. Circulating and local tissue level of AH was increased in severe congestive heart failure (CHF). However, the expression of ATI and AT2 in cardiac tissue with CHF remains controversial. Therefore, the present study was designed to investigate the protein expression of ATI and AT2 receptors in normal human myocardium and in human cardiac tissue with mild and severe CHF.

Human atrial tissues from normal subjects and CHF patients with ischemic cardiomyopathy and dilated cardiomyopathy were obtained from open-heart surgery and cardiac transplantation. ATI and AT2 receptor expression was investigated by immunohistochemical staining (IHCS). The results of IHCS was evaluated by IHCS staining density scores (0, no staining; 1, minimal staining; 2, mild staining; 3, moderate staining; and 4, strong staining).

Cardiac myocytes maintained in cell culture develop hypertrophy both in response to mechanical loading as well as to receptor-mediated signaling mechanisms. However, it has been shown that the hypertrophic response to these stimuli may be modulated through effects of intercellular contact achieved by maintaining cells at different plating densities. We recently developed the new special tissue culture system to culture minced cardiac tissue and to evaluate the apoptotic effect of different agent in cardiomyocytes. After 24 hours culture period, different culture conditions and factors may damage the cardiomyocyte. Therefore, we performed different methods to evaluate the viability of cardiomyocyte in cardiac tissue with 24 hours culture period in our special culture system.

Human cardiac atrial tissue was obtained from open-heart surgery (n=6). After cardiac tissue excision, the samples were immediately placed in oxygenated, nominally Ca2+-free Tyrode solution for transport to the laboratory. The time between excision and the beginning of laboratory processing was five to fifteen minutes.

C-type natriuretic peptide (CNP) is endothelial cell origin and has potent vasodilatory and antimitogenic actions. We reported that CNP was markedly increased in human cardiac tissue with severe congestive heart failure. To date, the effects of CNP on cardiomyocyte growth and death remain unclear. Therefore, the present study was designed to investigate the actions of CNP on apoptosis and apoptosis-related gene p53 expression in human cardiomyocytes.

Human cardiac atrial tissue was obtained from open-heart surgery (n=6). The cardiac tissue was minced and incubated in the special tissue culture system in the absence or presence of CNP (10-7 M) for 24 hours. These studies were repeated with HS-142-1 (HS, 10-6 M), a natriuretic peptide biological receptor antagonist. To detect the DNA fragmentation, in situ terminal deoxymucleotidyl transferase dUTP nick end labeling (TUNEL) was performed. The p53 expression was determined by immunohistochemical staining (IHCS). An average of 1000 nuclei was analyzed for TUNEL and p53 staining studies.

C-type natriuretic peptide (CNP) of endothelial cell origin via NPR-B receptor mediates antimitogenic and vasodilatory actions. As a circulating endocrine system, CNP plays a fundamental role in cardiorenal regulation. However, the actions of CNP on renal mesangial cell apoptosis remain poorly defined. Apoptosis might plays an important role during development of renal glomerular mesangial cells pathophysiology. The mechanisms of apoptosis include p53-dependent pathway and p53-independent pathway.

The hypothesis of this study is that CNP induces apoptosis through the process involving p53 gene in human glomerular mesangial cells via natriuretic peptide biological receptor. Therefore, the current study was designed to investigate the effects of CNP on apoptosis and p53 expression in human mesangial cell in the absence or presence of CNP biological receptor antagonist.

Cultured human mesangial cells (Clontech Lab., San Diego, CA) was incubated for 24 hours in the absence or presence of CNP (10-7 M). These studies were repeated with HS 142-1 (HS, 10-5 M), a CNP biological receptor antagonist.

Apoptosis or programmed cell death is involved in many diseases include end-stage renal failure. Apoptosis-related genes include both stimulate genes and inhibitory gene of apoptosis. The genes which stimulate apoptosis include p53 and p21-WAF. The genes which inhibit apoptosis include bcl-2 gene family. The mechanisms of apoptosis include p53-dependend pathway and p53- independent pathway. We hypothesized that apoptosis-related genes may activate in renal graft rejection after kidney transplantation. Therefore, the present study was designed to investigate apoptosis-related gene expression and localization by immunohistochemical staining (IHCS) in human renal tissues with graft rejection and compare with that in normal human renal tissue.

Human renal biopsy (n=5) were obtained after kidney transplantation with mild and moderate renal rejection. Normal human kidney biopsy was obtained during nephrectomy. P53, p21-WAF and Bcl-2 levels in renal tissue were determined by IHCS. The results of IHCS was evaluated by IHCS staining density scores (0, no staining; 1, minimal staining; 2, mild staining; 3, moderate staining; and 4, strong staining).

Regulation of apoptosis involves a number of genes that can be classified into broad categories. These include genes that act as effectors of apoptosis, such as p53, c-myc, bax, p21- WAF, and genes that primarily suppress apoptosis, such as Bcl-2 gene family. These genes have been reported to be responsible for the modulation of certain stress induced apoptosis and cell cycle arrest.

It has also been reported that apoptosis involved in cardiovascular diseases such as myocardial infarction, reperfusion injury, left ventricular hypertrophy, and hypertension. However, the expression of apoptosis-related genes in human cardiomyocytes in normal subjects and in patients with congestive heart failure (CHF) remains unclear. Therefore, the present study was designed to determine the expression and localization of apoptosis-related genes in human heart.

Five normal subjects and five end-stage CHF human ventricular cardiac tissues were obtained from cardiac transplantation. The expression of p53, p21-WAF and Bcl-2 were determined by immunohistochemical staining (IHCS).

Angiotensin II (AII) is a potent vasoconstrictor and mitogenic factor. The biological actions of All through AII receptors which include AT-1 and AT-2 receptors. However, the effects of AII on cardiomyocyte apoptosis and caspase expression remain controversial. Therefore, the current study was designed to investigate the actions of AII on human cardiomyocyte apoptosis and caspase-1 expression.

Human cardiac tissue was obtained from open-heart surgery (n=6). The cardiac tissue was minced and incubated in the special tissue culture system for 24 hours in the absence or presence of AII (10-7 M). These studies were repeated with losartan (10-6 M, AT-1 receptor antagonist) and PD123319 (PD, 10-6 M, AT-2 receptor antagonist). To detect the DNA fragmentation, TUNEL staining and DNA gel analyses were performed. The caspase-1 expression was determined by immunohistochemical staining (IHCS). An average of 1000 nuclei was analyzed for both TUNEL and caspase-1 IHCS studies.

Cyclosporine A (CsA) is the most effective and widely used immunosuppressant drug in heart, lung and kidney transplantation. However, the effect of CsA is limited by the significant toxicity. The mechanism of CsA-induced toxicity is remaining controversial. Cellular apoptosis is being suggested as a possible mediator of CsA toxicity. To date, regarding the effects of CsA on apoptosis and apoptosis-related gene regulation in cardiomyocytes remain unclear. Therefore, the current study was designed to investigate the effect of CsA on apoptosis and apoptosis-related gene p53 expression in human cardiomyocytes. We hypothesized that CsA induces apoptosis in human cardiomyocytes through p53-dependent pathway.

Human cardiac atrial tissue was obtained from open-heart surgery (n=5). The cardiac tissue was minced and incubated in the special tissue culture system for 24 hours in the absence or presence of CsA (10-7 M). To detect the DNA fragmentation, in situ terminal deoxymucleotidyl transferase dUTP nick end labeling (TUNEL) was performed.

Cyclosporine-A is the one of the most effective immunosuppressant drug in organ transplantation. However, cyclosporine-A has a potent cytotoxic effect. Cellular apoptosis is being suggested may contribute to the cyclosporine A-mediated cytotoxic action. To date, regarding the effects of cyclosporine-A on apoptosis and apoptosis-related genes expression in vascular smooth muscle cells remain unclear. Therefore, the current study was designed to investigate the actions of cyclosporine-A on apoptosis and p53 expression in human vascular smooth muscle cells.

Human saphenous vein was obtained from coronary artery bypass surgery (n=5). After saphenous vein removed, the tissue was minced and incubated in the special tissue culture system for 24 hours in the absence or presence of cyclosporine-A (10-6M). To detect the DNA fragmentation, in situ terminal deoxymucleotidyl transferase dUTP nick end labeling (TUNEL) was performed. The p53 level in human vascular smooth muscle cells was determined by immunohistochemical staining (IHCS). An average of 1000 nuclei was analyzed for both TUNEL and p53 EHCS studies.

Angiotensin II is a potent vasoconstrictor and mitogenic factor. However, the effects of angiotensin II on human vascular smooth muscle cells apoptosis remain controversial. Therefore, the current study was designed to investigate the actions of angiotensin II on human vascular smooth muscle cells apoptosis.

Human saphenous vein was obtained from coronary artery bypass surgery (n=6) and was minced and incubated in the special tissue culture system in the absence or presence of angiotensin II (10-6 to 10-12M) for 1, 2, 4, 8, 16, & 24 hours. These studies were repeated with losartan (10-6M, AT- 1 receptor antagonist) and PD-123319 (10-6M, AT-2 receptor antagonist). To detect the DNA fragmentation, in situ terminal deoxymucleotidyl transferase dUTP nick end labeling (TUNEL) and DNA agarose gel analyses were performed. An average of 1000 nuclei was analyzed for TUNEL studies.

TUNEL staining and DNA gel analysis demonstrated that angiotensin II increased apoptosis in human vascular smooth muscle cells.

Organisms are generally designated as “extreme” when they possess adaptations that allow them to flourish when one or more environmental parameters (e.g., temperature, ionizing radiation, oxygen, water, nutrients, inorganic ions) are at unusually high or low levels. A trade-off for this extraordinary resilience is that such organisms often grow poorly (if at all) in more conventional milieus. As a result, few of them are mammalian pathogens. (Anaerobic bacteria, which frequently thrive in the hypoxic conditions afforded by devitalized tissue, are an exception to this rule.)

A broader definition of “extremity” encompasses all organisms that occupy narrow or unusual environmental niches. In this more inclusive scheme, a wealth of viruses, bacteria, fungi, and parasites with pathogenic potential in humans can be considered extreme. These organisms often pose formidable diagnostic challenges. Many have extremely narrow host ranges, rendering passage in animals and development of model systems for infection difficult or impossible.

Opportunistic viral infections in immunosurpressed patients may morphologically involve only a few cells. Identifying an optimal area by LM is most important before an attempt at detection of viral particles by EM. The purpose of this communication is to emphasize unusual and important structural findings for some viruses.

BKV - BK polyomavirus latently infects renal tubular epithelial cells. Immunosuppressed patients many develop reactivation with resultant tubular cell damage and acute interstitial nephritis. Ultrastructural examination shows spherical and tubular virus particles of typical polyomavirus. Cross sections of the tubular form of this virus may contain electron dense cores (fig.l). Cells may be coinfected with HIV on rare occasions (fig.2).

JCV - JC polyomavirus is the major cause of progressive multifocal leukoencephalopathy (PML). JCV are rarely seen in reactive bizarre astrocytes. Biopsy of the tissue at the periphery of lesions of PML is likely to include JC-infected oligodendrocytes; it is important to appreciate that astrocytes, regardless of appearance, are unlikely to contain the virus.

“Emergence” of infectious disease agents in humans, domestic animals and wildlife during the past 20 years has been widely described. Perception of emergence derives largely from the application of improved identification methods, including refinements in molecular, serologic, and microscopy technologies that enable researchers to monitor species migration with greater sensitivity. The reality of emergence comes from shifts in genetic profiles and from influences of ecological changes often brought about by human interventions designed for economic or quality-of-life gains, and ecologic management. CDC has frequent involvement with many outbreaks caused by various infectious disease agents, some of which may be considered emerging. Notable unusual agents include filoviruses (Fig. 1) and hantaviruses (Fig. 2). More typically encountered agents include caliciviruses (Fig. 3) and influenza viruses (Fig. 4). Recently, threats of national and international bioterrorism have added to CDC's responsibilities for prompt identification of infectious agents.