Investigation of the function of Candida albicans Als3 by heterologous expression in Candida glabrata.

During hematogenously disseminated infection, blood-borne Candida albicans invades the endothelial cell lining of the vasculature to invade the deep tissues. Although the C. albicans Als3 invasin is critical for invasion and damage of endothelial cells in vitro, a C. albicans als3Δ/Δ mutant has normal virulence in the mouse model of disseminated infection. We hypothesized that the contribution of Als3 to virulence is obscured by the presence of additional C. albicans invasins. To elucidate the in vivo function of Als3, we heterologously expressed C. albicans ALS3 in Candida glabrata, a yeast that lacks a close ALS3 ortholog and has low virulence in mice. We found that following intravenous inoculation into mice, the ALS3-expressing strain preferentially trafficked to the brain, where it induced significantly elevated levels of myeloperoxidase, tumor necrosis factor, monocyte chemoattractant protein 1, and gamma interferon. Also, the ALS3-expressing strain had enhanced adherence to and invasion of human brain microvascular endothelial cells in vitro, demonstrating a potential mechanism for ALS3-mediated neurotropism. In addition, upon initiation of infection, the ALS3-expressing strain had increased trafficking to the cortex of the kidneys. With prolonged infection, this strain persisted in the kidneys at significantly higher levels than the control strain but did not induce an elevated inflammatory response. Finally, the ALS3-expressing strain had increased resistance to neutrophil killing in vitro. These results indicate that during disseminated infection, Als3 mediates initial trafficking to the brain and renal cortex and contributes to fungal persistence in the kidneys.

Uropathogenic Escherichia coli causes cortical tubular necrotic cell death and the release of macrophage migration inhibitory factor.

The macrophage migration inhibitory factor (MIF), a pro-inflammatory cytokine, is deregulated in acute kidney injury (AKI) through an unknown mechanism. In the present study, we used a previously described mouse model of ascending urinary tract infection in which uropathogenic Escherichia coli (UPEC) were transurethrally inoculated to induce kidney infections. Here, we show that urinary MIF was upregulated during AKI while MIF was abundantly expressed in the renal cortical tubules and that UPEC infection caused a decrease in tubular MIF. Infections with UPEC in vitro caused MIF release in a cell type-dependent manner, which was independent of receptor-mediated internalization, signal transduction, and transcription. Indeed, UPEC infection-induced necrotic cell death in vitro and in vivo correlated with extracellular acidification and processed MIF secretion. These data suggest that MIF is released by necrotic renal cortical tubular cells during UPEC infection.

The antimicrobial peptide cathelicidin protects the urinary tract against invasive bacterial infection.

The urinary tract functions in close proximity to the outside environment, yet must remain free of microbial colonization to avoid disease. The mechanisms for establishing an antimicrobial barrier in this area are not completely understood. Here, we describe the production and function of the cathelicidin antimicrobial peptides LL-37, its precursor hCAP-18 and its ortholog CRAMP in epithelial cells of human and mouse urinary tract, respectively. Bacterial contact with epithelial cells resulted in rapid production and secretion of the respective peptides, and in humans LL-37/hCAP-18 was released into urine. Epithelium-derived cathelicidin substantially contributed to the protection of the urinary tract against infection, as shown using CRAMP-deficient and neutrophil-depleted mice. In addition, clinical E. coli strains that were more resistant to LL-37 caused more severe urinary tract infections than did susceptible strains. Thus, cathelicidin seems to be a key factor in mucosal immunity of the urinary tract.

CD103+ CTL accumulate within the graft epithelium during clinical renal allograft rejection.

BACKGROUND: We have previously reported that activated CD8+TCRalphabeta+ cells that express high levels of the beta7 integrin CD103 (formerly alphaE, MLA) are present at the graft site during clinical renal allograft rejection. This observation potentially provides new insight into the mechanisms underlying renal allograft destruction because the ligand of CD103 is the epithelial cell-specific molecule E-cadherin, which is known to be expressed by critical graft functional elements such as the renal tubular epithelium. We herein used combined fluorescence-activated cell sorter (FACS) and immunohistochemical (IHC) analyses of transplant nephrectomy (TN) specimens to demonstrate that CD103+ cytolytic T lymphocytes (CTLs) specifically home to the graft epithelium during rejection episodes. METHODS: Serial sections of TN specimens undergoing histologically confirmed cellular rejection (n=7) were stained with anti-CD8 or anti-CD103 and were scored for the presence of positively stained cells within the tubular basement membrane. Freshly isolated graft-infiltrating lymphocytes were subjected to three-color FACS analyses to define the extended phenotypic characteristics of CD103+ cells detected by IHC. RESULTS: CD103+ cells in all specimens were biased towards an intratubular localization. On average, the percentage of CD103+ cells with an intraepithelial localization was 52.2+/-13.1 compared to 12.0+/-3.5 for pan CD8+ cells (mean+/-SE, n=5). FACS analyses confirmed that CD103+ cells detected by IHC exhibited the salient characteristics of CD8+ CTLs (large CD8+TCRalphabeta+CD62L-CD11a(hi)perforin+). The CD103- subset of graft-infiltrating CD8 cells also exhibited a CTL phenotype, but these were predominantly restricted to the graft interstitium. CONCLUSIONS: These data implicate CD103 as a homing receptor that targets graft-infiltrating CD8+ CTLs to the graft epithelium. Given the strong association of tubulitis with clinical rejection, these data are consistent with a role for the CD103+ CTL subset as an effector mechanism in renal allograft destruction.

Perinephric and intrarenal abscesses.

Perinephric and intrarenal abscesses remain a significant source of morbidity and mortality as well as a diagnostic dilemma. The history, epidemiology, disease classification, etiology, diagnosis, and treatment are reviewed, with special attention to new diagnostic and treatment modalities.

[The area of focal nephritis measured by echography: useful indications in patients with unexplained back pain in comparison with other assessments]. / Le aree di nefronia focale all'ecografia: utili indicazioni in pazienti con inspiegabili dolori dorsali e confronto con altre indagini.

In some patients undergoing an U.S. study of kidney for lumbodynia, it's sometime possible to visualize hypoechoic and areas poorly demarcated without distal acoustic enhancement, localized within the cortex and disrupting the cortico-medullary junction. These findings, called focal nephritis, associated to minimal retention of urine in the bladder, reflect an inflammatory process involving the renal parenchyma, in spite of normal urine analysis. To confirm this theory, 7 patients who presented these findings at US study of kidney underwent renal scintigram with labeled granulocytes. This test revealed the presence of focal bacterial nephritis in the same hypoechoic areas. Therefore US study of kidney combined with renal scintigram is useful to diagnose inflammatory process of the kidney in patients complaining lumbodynia.

Leptospirae of serotype lora of the serogroup Australis isolated for the first time from swine in the Netherlands.

Leptospirosis in swine, caused by serotype lora (serogroup Australis) was detected both serologically and by culture. The most important symptoms consisted in abortion during the final month of gestation and the birth of dead or not viable piglets.

Immune complex binding Streptococcus pyogenes type M12/emm12 in experimental glomerulonephritis.

In a rabbit model, we have previously reported evidence for a pathogenic role of streptococcal IgG Fc-binding proteins (IgGFcBP) in poststreptococcal glomerulonephritis (PSGN). These proteins, of the M protein family, were shown to trigger anti-IgG production and enhance renal deposition of IgG and/or immune complexes (ICs), with resulting activation of complement and cytokine cascades. In the present study, type M12/emm12, group A streptococci (GAS) were found often to bind artificial ICs, viz. peroxidase-anti-peroxidase rabbit IgG (PAP) or tetanus toxoid-anti-tetanus human IgG (TAT), rather than monomeric IgG. Animals injected with each of four IC binding clinical isolates (from patients with scarlet fever or PSGN) showed pronounced inflammatory and degenerative glomerular changes, morphologically similar to human PSGN, with membrane thickening and IgG and complement C3 deposition, as well as secretion of IL-6 and TNF-α by mesangial and endothelial cells. In contrast, non-binding strains (two from asymptomatic carriers and one from a PSGN case) failed to trigger any renal changes. Only the IC binding strains induced elevated titres of anti-IgG. Though the streptococcal binding component(s) has not been demonstrated, the selective binding of ICs by type M12/emm12 strains appears important for the well-known, marked nephritogenic potential of this GAS type.

Survey of rats (Rattus norvegicus) in Kuwait for the presence of Leptospira.

A survey of brown rats (Rattus norvegicus) was made in Kuwait in 1979 for the presence of Leptospira organisms. Kidney tissue from 49 rats, trapped mostly from various Kuwait City districts, were cultured on E.M.J.H. and Stuart media. Eight leptospira strains were isolated; all strains were identical and belonged to Leptospira interrogans Canicola serogroup; they were later identified as a new serovar kuwait. The frequency of L. interrogans group Canicola carriers among the local rats in Kuwait was 16.3%, which is higher than reported so far. A summary of published data on the isolation of Leptospira other than L. interrogans serogroup Icterohaemorrhagiae from rats, particularly the brown rat, is presented with emphasis on the Middle East.

Cytomegalovirus replicates efficiently in human kidney mesangial cells.

Human cytomegalovirus (HCMV) is a major renal pathogen in congenitally infected infants and renal allograft recipients. We postulated that a specific renal cell type was involved in HCMV infection and reactivation. Human fetal kidney cortex cell cultures were assayed for their ability to support HCMV infection. Infectious center assays indicated that the low level of viral replication observed by virus yield assay occurred from a fraction of the cells in the mixed cultures. Virus-specific immunofluorescence and in situ hybridization documented the presence of HCMV-specific protein and nucleic acid, respectively, in a morphologically distinct cell type. These cells were purified, were identified as kidney mesangial cells, and were observed to support efficient HCMV replication. Our research identifies mesangial cells as a renal cell type that supports HCMV replication and provides evidence to implicate these cells in the pathogenesis of HCMV-induced renal disease.

Susceptibility of a line of dolphin kidney cell culture to several herpesviruses.

A cell line was established from cell cultures of kidney cortex of a pantropical spotted dolphin, Stenella attenuate. The replication of 6 strains of herpesviruses was studied in the cells. Five strains of them, herpes simplex virus type I and type II, equine rhinopneumonitis virus, infectious bovine rhinotracheitis virus and Aujeszky's disease virus, were grown fairly well in showing clear cytopathic effects and plaques under agar overlay medium.

Viral susceptibility of an established cell line of swine embryo kidney.

The viral susceptibility of a cell line, named KSEK6, newly established from the kidney cortex of swine embryo was tested for the indication of CPE occurrences and also plaque formations. The multiplication of porcine adenoviruses was considerably high in the cells among the virus strains tested though plaques of these viruses were hardly visible under agar overlay medium. Two strains of swine enteroviruses, Aujeszky's disease virus and hemagglutinating encephalomyelitis virus also multiplied well in a similar order to those received in the other cells employed.

Multifunctional nature of P fimbriae of uropathogenic Escherichia coli: mutations in fsoE and fsoF influence fimbrial binding to renal tubuli and immobilized fibronectin.

P fimbriae of the F7(1) serotype of Escherichia coli are composed of a major subunit, FsoA, and of three minor proteins named FsoG, FsoE, and FsoF. FsoG is the Gal alpha(1-4)Gal-specific lectin. We assessed mutated recombinant strains each deficient in one fimbrial component for adhesion to frozen sections of rat cortical kidney and to fibronectin immobilized on glass. Rat kidney lacks the Gal alpha(1-4)Gal-containing glycolipids. The fsoG mutant strain was as adhesive to sections of rat kidney and to fibronectin-coated glass as was the recombinant strain expressing the complete fso gene cluster. The fsoA mutant strain was highly adhesive to fibronectin and to kidney sections. In the rat kidney, the adhesion of these strains was predominantly localized to sites of basolateral membranes of tubuli. The fsoE and the fsoF mutant strains were slightly less adhesive to kidney structures and failed to adhere to fibronectin. The fsoE, fsoF double mutant strain adhered neither to fibronectin nor to kidney sections. None of the fso recombinant strains reacted with soluble fibronectin, suggesting that the interaction is dependent on the conformation of the fibronectin molecules. Recombinant strains expressing the F7(2), F8, F11, F13, and F14 serovariants of the P fimbria also showed adherence to immobilized fibronectin. The results show that in addition to binding to globoseries of glycolipids via the G protein, the P fimbriae of uropathogenic E. coli exhibit a tissue-binding property influenced by fsoE and fsoF gene products and with affinity for basolateral membranes and fibronectin.

Acute human pyelonephritis: leukocytic infiltration of tubules and localization of bacteria.

The fine structural details of how leukocytes appear in the lumen of tubules and the localization of bacteria in the tubulo-interstitial space were studied by light and electronmicroscopy in renal cortical biopsy specimens from three patients with acute pyelonephritis. The cells of interstitial infiltrates infiltrated and sometimes disrupted the cortical collecting tubules preferentially, while inflammatory infiltration of the proximal and distal convoluted tubules occurred more rarely. Since the emigration of tubular wall-localized individual leukocytes into the lumen was not observed even in long series of thin sections, focal inflammatory disruption of the uriniferous ducts was considered to be the morphological basis of the intratubular accumulation of leukocytes. The structural simplicity of the collecting tubular cells is suggested to be the reason for their preferential involvement in the drainage of the interstitial suppuration, although a role for specific carbohydrate receptors cannot be excluded. The bacteria were usually found within the neutrophilic granulocytes and macrophages of the interstitial infiltrates, and within and among the cells of leukocyte casts. Additionally, pure bacterial colonies were noticed in the lumen of a few collecting tubules. The problem of the adherence of the bacteria to the surface of the tubular cells is discussed.

Progressive multifocal leucoencephalopathy: detection of papovavirus JC in kidney tissue.

Cellular DNA of the kidney from a patient with PML was analyzed by reassociation kinetics for the presence of JC virus DNA. Various amounts of viral DNA sequences were detected in different areas of the kidney. The highest concentration (175 genome equivalents/cell) was found in the renal medulla and there were almost none in the renal cortex. Differentiation from the closely related BK virus was carried out by reassociation kinetics and restriction enzyme cleavage with subsequent Southern blot analysis. The enzyme Hind II, which does not cleave within the BK virus genome, generated four restriction enzyme fragments in the cellular DNA from the kidney, thus documenting the presence of JC virus DNA. By examination of the renal DNA with the "no-cut" restriction enzyme XHO I and the "one-cut" enzymes Eco RI and BAM HI it was possible to show that free and not integrated viral DNA was present in these cells. Nonhomogeneous defective DNA bands were not detectable. By in situ hybridization the epithelial cells lining the collecting tubules were found as predominant site of the viral infection in the kidney.

Nuclear and cytoplasmic inclusions in disseminated human cytomegalovirus infection.

A case of disseminated human cytomegalovirus infection is described in a full-term female who expired 3 1/2 h after birth. Cytomegalic inclusions, both intranuclear and intracytoplasmic, were observed mainly in the kidneys, liver, lungs, and anterior pituitary but were not seen in the bone marrow and spleen. Whereas the nuclear inclusions consisted of an amorphous filamentous meshwork with a variable number of pleomorphic capsids, the cytoplasmic inclusions were composed of membrane-bound aggregates of mature virions with dense cores and multilayered envelopes. The complex envelopment process of cytomegalovirus appears to involve successive coats derived from the nuclear membrane and from the endoplasmic reticulum or cytoplasmic vesicles.

Host-parasite interactions in the pathogenesis of experimental renal candidiasis.

To study the pathogenesis of renal candidiasis, viable Candida albicans blastospores were injected directly into the left renal artery of New Zealand white rabbits. The progression of the disease was followed by light and electron microscopy over a 6-day period. Within 5 minutes after injection of the yeasts, the organisms localized within glomerular and peritubular capillaries of the cortex. Localization of yeasts within the capillaries occurred through adherence, demonstrated by the presence of surface fibrils originating from the yeast cells. Two to 10 hours later, inflammatory nodules comprised of polymorphonuclear leukocytes formed within capillary lumina. Many of the entrapped yeasts remained viable and extended through adjacent endothelium and epithelium by the formation of germ tubes which penetrated between or directly through intact host cells. After 24 hours, numerous hyphal forms were observed within tubules of the cortex, and some necrotic host cells were noted at sites of penetration. Abscesses replaced renal parenchyma in focal areas during subsequent time intervals. These studies indicate that attachment of Candida albicans to endothelium within capillaries of the cortex is a key event in the disease process. Also, growth of germ tubes into renal tubules provides a temporal advantage for amplification of Candida organisms.