Bone finds and their medicolegal examination: a study from Hesse, Germany.

Bones found by chance can be of great criminal or historical interest. The nature of their appraisal depends on the individual case, the locally effective legislation and the available resources. To assess whether a find is relevant with respect to criminal investigation, the circumstances of the find and the results of the forensic examination carried out by trained personnel must be considered. The aim of this study was to obtain an overview of the circumstances and nature of the finds as well as the results of the subsequent expert opinions by evaluating bone finds from the federal state of Hesse, Germany. For this purpose, over a 10-year period from 2011 to 2020, all bone finds examined at the Institutes of Legal Medicine in Gießen and Frankfurt am Main, Germany, were evaluated retrospectively with regard to the locations and circumstances of the finds, their nature (human or non-human), the postmortem interval, possible traces of violent impact and the results of further examinations. Of the 288 bone finds evaluated, 38.2% were found in forests, meadows and parks. In 50.7%, the finds contained human bones, of which 37.0% had a forensically relevant postmortem interval of 50 years or less. Evidence of trauma was described in 77.4% of the human bone cases: postmortem damage in 78.8%, peri-mortem injury in 9.7% and ante-mortem injury in 11.5%. DNA examinations were performed in 40.4% of the human bone finds. They yielded STR profiles in 81.3%, leading to a definite identification in 35.4%. Among the non-human bones sent in, the most common were bones from pigs (23.4%), deer (18.1%), cattle (16.4%), roe deer (11.7%) and sheep (11.7%). The macroscopic examination is the first step of the forensic-osteological evaluation and sets the course for further examinations or investigations. DNA examinations are of great importance for the reliable identification of human bones. They were responsible for 70.8% of successful identifications.

[A survey of preoperative perforator mapping in perforator flap surgery]. / Umfrage zu Methoden der präoperativen Lokalisation von Perforans-Gefässen zur erleichterten Planung von Perforans-Lappenplastiken.

INTRODUCTION: The frequency of perforator flap surgery is increasing. As such, the claim to identify and localize the perforators is closely related to preoperative perforator imaging. Starting with Doppler sonography magnetic resonance imaging (MRI) followed early in the 90s. Recently, three- and four-dimensional computer tomographic angiography and venography present impressive preoperative illustrations of perforators. HYPOTHESIS: Duplex ultrasound dominates in clinical practice for perforator imaging. METHODS: We performed a survey using a multiple choice questionnaire which was distributed among consultants of the German plastic surgery society (DGPRAC). Response rate was 46% with 132 institutions responding. RESULTS: Preoperative perforator imaging is applied in 77%. Duplex sonography is predominant with 70%, followed by colour Doppler ultrasound (42%). CT angiography (7%) and MRI angiography (8%) are currently underutilized. The amount of perforator flaps performed did not favor any preoperative perforator diagnostic. However, among those performing more than 30 perforator flaps per year, only 2% did not perform any perforator diagnostic at all. CONCLUSION: Preoperative perforator imaging is applied in 3/4 of all perforator flaps with duplex ultrasound dominating. Angio-CT and Angio-MRI are currently infrequently used for preoperative perforator imaging in Germany to date.

[Nicotine in plastic surgery : a review]. / Nikotinkonsum und plastische Chirurgie : Eine aktuelle Ubersicht.

BACKGROUND: The surgical complication rate is significantly increased in active smoking patients. However there are no evidence-based recommendations regarding smoking among patients seeking plastic surgical procedures. METHODS: MEDLINE analysis was performed of all relevant clinical and experimental papers from 1965 to 2008. RESULTS: In face-lift operations smokers present a 13-fold risk of skin necrosis. In mamma reduction procedures the risk among smokers is doubled for number of complications, with T-incision site necrosis (odds ratio 3.1) and infection rate (OR 3.3) significantly elevated among active smokers. Transverse rectus abdominis myocutaneous flaps for breast reconstruction are associated with significantly higher flap necrosis rates for smokers than nonsmokers (19% vs 9%, P=0.005). The smoking history can be indicative, but usually the number of cigarettes is drastically underestimated. Cotinine testing is a method of determining smoking quantitatively up to 4 days before testing. CONCLUSION: Four weeks of abstinence from smoking reduces smoking-associated complications. Despite the published evidence, smoking is no longer relevant in the German 2008 Disease-Related Group for plastic surgical procedures.

Inhibition of tubule-cell proliferation to prevent cyst formation: a new avenue to treat ADPKD?

Tubule-cell hyperproliferation precedes cyst development in autosomal dominant polycystic kidney disease (ADPKD). Parker et al. report that insulin-like growth factor-1 stimulates ADPKD cell proliferation by activating Ras and Raf; inhibition of Ras or Raf abolished this effect. Inhibiting tubule-cell proliferation could halt cyst formation and prolong survival of functional tubules, offering a new treatment for ADPKD patients.

Evidence of angiogenesis and microvascular regression in autosomal-dominant polycystic kidney disease kidneys: a corrosion cast study.

Autosomal-dominant polycystic kidney disease (ADPKD) accounts for about 10% of all cases of chronic renal failure requiring dialysis. The disease is characterized by proliferation of renal epithelial cells and formation of cysts that expand over years and replace the normal parenchyma of the kidney. As the cysts grow, the volume of the kidney can increase by more than 10-fold, implying that remodeling and expansion of the vasculature must occur to provide oxygenation and nutrition to the cyst cells. Our previous studies support the notion that there is angiogenesis in ADPKD. We report here results from resin casting of ADPKD kidneys vasculature. In this study, the corrosion-casting method was used in conjunction with scanning electron microscopy to study the vascular architecture and the evidence for angiogenesis in ADPKD kidneys. We found a well-defined vascular network around the cysts forming a 'vascular capsule' somewhat similar to that described in avascular leiomyomata. We also found that the normal vascular architecture is lost and replaced by an assortment of capillaries of larger size than those in the normal kidney, mixed with flattened and spiral arterioles, damaged glomeruli, and atresic venules, indicative of regression of the microvasculature. In the same areas, there was capillary sprouting, considered the hallmark of angiogenesis. The present study documents regression changes of the vasculature and confirms the existence of angiogenesis in ADPKD kidneys, and suggests the use of inhibitors of angiogenesis as a possible avenue for the treatment of the disease.

Information access at the point of care: what can we learn for designing a mobile CPR system?

OBJECTIVES: Hospitals have started to migrate their paper-based records to computerized patient records (CPR). The majority of today's CPR systems are stationary, which means that physicians use a clinical workstation to access CPR information. But health care professionals need to request and enter information at different locations, for example, on their daily ward round. This suggests the use of mobile computers, enabling an ubiquitous access to needed data. Different studies show that health care professionals are reluctant to use poorly designed mobile CPR systems, as the work at the point of care is very time-pressured and hectic. To design a system with high acceptance, it is essential to obtain empirical insight into the work practices and context in which the mobile CPR system will be used. METHOD: We investigated the physicians' work with the patient record during their daily round. With the help of a compact notation method, the physicians' interaction with the information system was recorded in real time. Fourteen physicians from three different departments (internal medicine, surgery, and geriatrics) of a middle-sized Swiss hospital participated in our study. RESULTS: Physicians have clear access preferences when they interact with the patient record during their daily round. There exists a clear profile of access frequencies and patterns, respectively. As an example, approximately 50% of all patient record accesses concern information about medications, vital signs and lab test results. DISCUSSION/CONCLUSION: A CPR system which is designed to reflect the access frequencies and patterns should improve the efficiency of data entry and retrieval and thus result in a system with high acceptance among physicians in the demanding environment during hospital rounds.

Do connexin 43 gap-junctional hemichannels activate and cause cell damage during ATP depletion of renal-tubule cells?

UNLABELLED: We review our evidence in favour of the hypothesis that gap-junctional hemichannels (GJH) are activated by depletion of adenosine triphosphate (ATP) in human renal proximal tubule cells in primary culture (hPT cells). Undocked GJH permit fluxes of ions and hydrophilic molecules up to 1 kDa, and thus their opening can cause alterations of cell composition conducive to cell damage. We show that hPT cells express connexin 43 (Cx43) (at the mRNA and protein levels) and that the protein is expressed on the plasma membrane. Moderate levels of pharmacological depletion of ATP increased plasma-membrane permeability, as shown by loading with the hydrophilic dye 5/6 carboxyfluorescein (CF, 376 Da) and other low-molecular weight dyes, but not with fluorescein-labelled dextran (>1500 Da). Roles for endocytosis and activation of purinergic-receptor channels were experimentally ruled out. Moderate ATP depletion also caused necrosis, assessed by cell permeabilization to propidium iodide. Prolonged exposure to gadolinium reduced both the dye loading and the necrosis induced by ATP depletion, i.e. it protected the cells. Cx43 overexpressed in insect cells, purified to homogeneity and reconstituted in proteoliposomes formed hemichannels that are activated by dephosphorylation of Ser368, a residue in a protein-kinase-C consensus phosphorylation sequence near the end of the C-terminal domain. CONCLUSIONS: (1) ATP depletion of hPT cells induces a Gd3+-sensitive permeability of the plasma membrane to hydrophilic dyes with a cut-off size consistent with Cx43 GJH. (2) ATP depletion also increases the percentage of necrotic cells, an effect also reduced by Gd3+. (3) The experiments with purified Cx43 reconstituted in liposomes suggest that dephosphorylation of Ser368 is sufficient to activate GJH, although other mechanisms may be involved in some cells.

Gap-junctional hemichannels are activated by ATP depletion in human renal proximal tubule cells.

We present evidence suggesting that gap-junctional hemichannels (GJH) may be involved in acute ischemic injury of human renal proximal tubule cells (hPT cells). Two GJH, from neighboring cells, join to form an intercellular gap junction channel (GJC). Undocked GJH are permeable to hydrophilic molecules up to 1 kDa, and their opening can significantly alter cell homeostasis. Both GJC and GJH formed by connexin 43 (Cx43) are activated by dephosphorylation. Hence, we tested whether GJH activation during ATP depletion contributes to cell damage in renal ischemia. We found that hPT cells in primary culture express Cx43 (RT-PCR and Western-blot analysis) at the plasma membrane region (immunofluorescence). Divalent-cation removal or pharmacological ATP depletion increased cell loading with the hydrophilic dye 5/6 carboxy-fluorescein (CF, 376 Da) but not with fluorescein-labeled dextran (>1500 Da). Endocytosis and activation of P2X channels were experimentally ruled out. Several GJC blockers inhibited the loading elicited by PKC inhibition. Double labeling (CF and propidium iodide) showed that both Ca(2+) removal and ATP depletion increase the percentage of necrotic cells. Gadolinium reduced both the loading and the degree of necrosis during divalent-cation removal or ATP depletion. In conclusion, GJH activation may play an important role in the damage of human renal proximal tubule cells during ATP depletion. These studies are the first to provide evidence supporting a role of GJH in causing injury in epithelial cells in general and in renal-tubule cells in particular.

Differential regulation of interleukin-10 production by genetic and environmental factors--a twin study.

Interleukin-10 (IL-10) has a critical role in the regulation of immune responses. The relative contribution of genetic and environmental factors to IL-10 production is under debate. We performed a twin study in 246 monozygotic and dizygotic twins to assess the heritability of IL-10 production after LPS stimulation in whole blood. In addition, the influence of promoter single nucleotide polymorphisms (-1082, -819 and -592) on transcriptional activity and their binding to nuclear factors was studied in luciferase reporter gene and electrophoretic mobility shift assays. IL-10 production showed a genetic determination with a heritability of 0.5. Decreasing body mass index (BMI), smoking and female gender lead to decreased IL-10 production. In monocytes, the -1082A allele showed higher binding affinity to the transcription factor PU.1 resulting in decreased transcriptional activity of -1082A promoter haplotypes. Genetic determination of IL-10 secretion is probably lower than that previously reported. Fifty percent of the observed variability explained by genetic factors. Female individuals produce less IL-10 than male subjects. Environmental factors like smoking and decreasing BMI exert suppressing effects on IL-10 production. Although the -1082A allele shows higher binding affinity to the PU.1 transcription factor and lower transcriptional activity, this polymorphism probably explains only a small fraction of the observed heritability.

Differential association of polymorphisms in the TNFalpha region with psoriatic arthritis but not psoriasis.

OBJECTIVE: To investigate the potential association of tumour necrosis factor alpha (TNFalpha) microsatellite and promoter alleles with psoriatic arthritis (PsA). METHODS: DNA from 89 white patients with PsA, 65 patients with psoriasis, and 99 healthy white controls was investigated for two TNFalpha promoter (-238 and -308) and three microsatellite polymorphisms (TNFa, c, and d). Patients had previously been studied by serology for HLA class I antigens and by sequence-specific polymerase chain reaction for DRB1* alleles. In addition, TNFalpha production of Ficoll separated peripheral blood mononuclear cells (PBMC) into culture supernatants after stimulation with lipopolysaccharide, alphaCD3 antibodies, phytohaemagglutinin, and streptococcal superantigen C was determined. RESULTS: A significant, HLA class I independent increase of the TNFa6c1d3 haplotype was found in the group with PsA but not among patients with psoriasis (32% v. 8%, pc<0.008; relative risk (RR)=5.3). In addition, patients with PsA showed a marked decrease of the TNF308A promoter allele (6% v. 18%; pc<0.008; RR=3.5) compared with healthy controls, which was independent of the increased frequency of the -238A polymorphism in this group. PBMC from patients with PsA secreted significantly less TNFalpha than cells from patients without arthritis. In particular, the TNFa6 microsatellite was associated with decreased TNFalpha production. CONCLUSION: These data indicate that allelic variations at the TNFalpha locus influence susceptibility to PsA. Decreased production of TNFalpha is at least in part genetically determined and might be related to the development of arthritis. However, the association of the TNF308G allele with the disease also points to other disease related haplotypes with still unknown susceptibility genes.

Atypical glandular cells of undetermined significance. Subtyping as a predictor of outcome.

OBJECTIVE: To determine whether subtyping of atypical glandular cells of undetermined significance (AGUS) cervical smears into reactive process favored (AGUS-RPF) and not otherwise specified (AGUS-NOS) correlates with rates of underlying pathology. STUDY DESIGN: We performed a retrospective chart review of 129,676 routine Pap smears at Kaiser Permanente, San Diego, from June 1998 to November 1999. One hundred fifty of the 129,676 were evaluated as AGUS (0.12%). Subjects with concomitant AGUS/SIL Pap smears, a prior history surgery for dysplasia, prior AGUS, a history of cancer or prior hysterectomy were excluded from the study. The remaining AGUS smears were then subtyped into AGUS-NOS and AGUS-RPF based on the criteria of the 2nd Workshop of the Bethesda System. Sixty-eight subjects with an initial AGUS smear underwent evaluation with colposcopy, endocervical curettage, endometrial biopsy and directed cervical biopsies. Significant pathology was determined to be any tissue diagnosis that required further treatment more than a follow-up Pap smear. AGUS subclassifications and underlying pathology were then compared using the chi 2 test with Fisher's Exact Test. RESULTS: Twenty-seven patients (40%) had AGUS-NOS, and 41 (60%) had AGUS-RPF. There were no significant differences between the groups in regard to age, race, parity, menopause status, HRT use or tobacco use. CONCLUSION: Subtyping AGUS cervical smears correlates with underlying rates of pathology. However, AGUS-RPF smears were still associated with an approximately 10% incidence of significant underlying pathology. Patients with AGUS on cervical smears need thorough evaluation, regardless of subtyping.

Angiogenesis in autosomal-dominant polycystic kidney disease.

BACKGROUND: Autosomal-dominant polycystic kidney disease (ADPKD) is a genetic disorder that is responsible for approximately 10% of all cases of end-stage renal disease (ESRD). It is characterized by the formation of epithelial cell cysts, an increase in the extracellullar matrix, and vascular alterations believed to be the result of compression by the cysts. Our recent observations demonstrated a rich vascular network on the surface of the cysts, and thus, we postulated that angiogenesis could be a factor in the progression of ADPKD. METHODS: Kidneys removed from patients with ADPKD were studied using (1) angiographs, (2) immunostaining [factor VIII-related antigen, vascular endothelial growth factor (VEGF), VEGF receptors 1 and 2 (VEGFR-1 and VEGFR-2), metalloproteinase-2 (MMP-2), and integrin alphavbeta3], and (3) Western blot analysis and enzyme-linked immunosorbent assay. The expression of VEGF165 in ADPKD cells in culture was determined. RESULTS: There was (1) an extensive capillary network in the cyst wall of ADPKD kidneys, (2) morphological evidence of vascular malformations, (3) expression of VEGF165 in cyst cells of VEGFR-2 in endothelial cells and an absence of VEGFR-1 in endothelial cells, (4) secretion of VEGF165 by ADPKD cyst cells in culture, and (5) coexpression of matrix MMP-2 and integrin alphavbeta3 in vessels from ADPKD. CONCLUSIONS: There is angiogenesis in ADPKD. This process may be necessary for cyst cells to grow and may be responsible for increased vascular permeability facilitating fluid secretion into the cysts. Neovascularization may result in the formation of aneurysms responsible for the renal bleeding in this disease.

IL10.G microsatellites mark promoter haplotypes associated with protection against the development of reactive arthritis in Finnish patients.

OBJECTIVE: To investigate the association of microsatellites and single-nucleotide promoter polymorphisms (SNPs) in the gene for the cytokine interleukin-10 (IL-10) with susceptibility to and outcome of reactive arthritis (ReA). METHODS: From genomic DNA, IL-10 microsatellites G and R and IL-10 promoter polymorphisms at positions -1087 and -524 were typed by polymerase chain reaction, automated fragment length analysis, and restriction fragment digestion in 85 Finnish patients with ReA and 62 HLA-B27-positive Finnish controls. ReA patients had been followed up for 20 years. Genotypes and haplotypes of IL-10 were correlated with distinct features of the disease course, such as triggering agent, chronic arthritis, development of ankylosing spondylitis, and other chronic features. RESULTS: There was a significant decrease in the promoter alleles G12 (allele frequency 0.206 versus 0.033; corrected P < 0.001, odds ratio 0.14) and G10 (0.183 versus 0.092; P < 0.05, odds ratio 0.44) in the ReA group compared with the HLA-B27-positive controls. Chronic arthritis developed significantly more frequently in the B27-positive subjects than in the B27-negative subjects (P < 0.05) as well as in patients with [corrected] the IL10.G8 allele. No associations were observed for either SNP or for the IL10.R microsatellite polymorphism. CONCLUSION: IL10.G12 and G10 microsatellite alleles show a strong protective effect against the development of ReA in Finnish subjects. Since these polymorphic markers themselves do not have direct functional implications, they most likely mark promoter haplotypes with distinct functional properties, suggesting that differential production of IL-10 is an important susceptibility factor for the development of ReA.

Different transcriptional activity and in vitro TNF-alpha production in psoriasis patients carrying the TNF-alpha 238A promoter polymorphism.

Genes encoded on chromosome 6 within the major histocompatibility complex region are thought to play an important role in the pathogenesis of psoriasis. A potential candidate gene is tumor necrosis factor alpha. The tumor necrosis factor alpha promoter contains several polymorphisms including two G-->A transitions at position -308 and -238, which are the most common in Caucasian populations. The TNF238.2 (-238A) allele has been strongly associated with psoriasis. We have investigated the effect of the -238 and -308 variants on transcription of the tumor necrosis factor alpha gene in luciferase reporter gene assays. In addition, peripheral blood mononuclear cells of 47 patients with psoriasis and 43 controls were stimulated with different antigens and mitogens (streptococcal sonicate and superantigen, lipopolysaccharide, phorbol-12-myristate, phytohemagglutinin, CD3 antibodies) and tumor necrosis factor alpha production was measured in supernatants by enzyme-linked immunosorbent assay. The psoriasis-associated tumor necrosis factor alpha promoter allele TNF238.2 showed a significantly decreased transcriptional activity. Peripheral blood mononuclear cells carrying this allele produced significantly less tumor necrosis factor alpha after stimulation with T cell mitogens and streptococcal antigens in comparison to controls. The promoter allele TNF238.2 seems to influence tumor necrosis factor alpha production; a possible role in the pathogenesis of psoriasis has to be further evaluated.

Role of multidrug resistance P-glycoprotein in the secretion of aldosterone by human adrenal NCI-H295 cells.

We determined the role of the multidrug resistance (MDR1) gene product, P-glycoprotein (PGP), in the secretion of aldosterone by the adrenal cell line NCI-H295. Aldosterone secretion is significantly decreased by the PGP inhibitors verapamil, cyclosporin A (CSA), PSC-833, and vinblastine. Aldosterone inhibits the efflux of the PGP substrate rhodamine 123 from NCI-H295 cells and from human mesangial cells (expressing PGP). CSA, verapamil, and the monoclonal antibody UIC2 significantly decreased the efflux of fluorescein-labeled (FL)-aldosterone microinjected into NCI-H295 cells. In MCF-7/VP cells, expressing multidrug resistance-associated protein (MRP) but not PGP, and in the parental cell line MCF7 (expressing no MRP and no PGP), the efflux of microinjected FL-aldosterone was slow. In BC19/3 cells (MCF7 cells transfected with MDR1), the efflux of FL-aldosterone was rapid and it was inhibited by verapamil, indicating that transfection with MDR1 cDNA confers the ability to transport FL-aldosterone. These results strongly indicate that PGP plays a role in the secretion of aldosterone by NCI-H295 cells and in other cells expressing MDR1, including normal adrenal cells.

Secretion of platelet-activating factor is mediated by MDR1 P-glycoprotein in cultured human mesangial cells.

MDR1 P-glycoprotein (Pgp), the product of the MDR1 gene involved in multidrug resistance in cancer cells, is also expressed in normal tissues. In the human kidney, it is localized in the mesangium, the proximal tubule, the thick ascending limb of Henle's loop, and the collecting duct. Pgp actively transports lipophilic xenobiotics, peptides, steroids, and lipids, and perhaps endogenous substrates. It has been shown previously that human mesangial cells in culture express active Pgp and that the expression of Pgp can be down-regulated by exposure to antisense oligonucleotides. Mesangial cells do not express multidrug resistance-related protein (MRP). Experiments were performed to determine whether 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine (generically platelet-activating factor, PAF) is a substrate of Pgp in human mesangial cells in culture. This study found: (1) PAF C-16 and analogs inhibited Pgp-mediated efflux of rhodamine 123 by 59 to 88% in multidrug-resistant KBV-1 cells and by 85 to 97% in cultured human mesangial cells. (2) In mesangial cells stimulated with A23187, the secretion of endogenously produced PAF was inhibited by >80% by the Pgp blockers verapamil, cyclosporin A, PSC-833, vinblastine, and adriamycin. (3) Preincubation with MDR1 antisense oligonucleotides also blocked PAF secretion by human mesangial cells. PAF analogs do not modify the transport of MRP substrates in MCF-7/VP cells expressing MRP but not Pgp. These results indicate that PAF is an endogenous substrate of Pgp in human mesangial cells. Inhibition of Pgp transport may be useful in reducing glomerular damage occurring in pathologic conditions where PAF secretion is elevated.

P-glycoprotein functions and substrates: possible roles of MDR1 gene in the kidney.

There is a renewed attention on the multidrug resistance genes and their products, P-glycoproteins, since recent molecular and functional studies revealed unexpected functions in normal tissues. There are two types of human P-glycoprotein: Type I, encoded by the MDR1 gene, present in excretory organs and in non-polarized cells; and Type II, encoded by MDR2, present in the canalicular membrane of hepatocytes. MDR1 Pgp transports xenobiotics, peptides, steroids, and phospholipids, and is also a regulator of swelling-activated chloride channels. MDR2 Pgp is exclusively a phosphatidylcholine translocase. In the kidney, the MDR1 gene and protein are expressed in mesangial, proximal tubule, thick loop of Henle, and collecting duct cells. In mesangial and proximal tubule cells Pgp transports xenobiotics. Concomitant exposure of kidney cells to two Pgp substrates results in increased cell toxicity. Extracts from supernatants of mesangial cell cultures inhibit Pgp-mediated transport, suggesting that a mesangial-cell metabolite could be a substrate of Pgp. Active vitamin D3 and platelet activating factor inhibit Pgp transport and are possible endogenous substrates in proximal tubule and mesangial cells, respectively. Pgp could be also a regulator of swelling-activated chloride channels present in the kidney.

Expression of MDR1 (multidrug resistance) gene and its protein in normal human kidney.

P-glycoprotein (Pgp), the product of the multidrug resistance (MDR) gene overexpressed in cancer cells, is present also in normal tissues. In the kidney, MDR1 Pgp has been found in the proximal tubule and in cultured mesangial cells. In situ hybridization and immunohistochemistry were used to determine the complete nephronal localization of MDR mRNA and its product, Pgp, in the human kidney. MDR mRNA expression was studied with the use of nonradioactive in situ MDR RNA probes. MDR1 Pgp was immunolocalized using the specific monoclonal antibody MRK16. The presence of MDR mRNA was confirmed in proximal tubules and mesangium, and demonstrated as well in thick limb of Henle's loops and in collecting ducts. MDR1 Pgp colocalized in the same nephronal segments. This suggests that, in addition to secreting xenobiotics, Pgp may play a role in the transport of endogenous substrates or in the regulation of Cl- channels.

Xenobiotic transport differences in mouse mesangial cell clones expressing mdr1 and mdr3.

P-glycoprotein (PGP), which confers multidrug resistance to cancer cells, is expressed in mouse kidney proximal tubule and mesangium. We report on the expression of PGP and its xenobiotic transport function in mesangial cells. Studies were performed in a mouse mesangial cell line (TKGM) and two cell clones. Ribonuclease protection assay and Western blot analysis demonstrated that TKGM cells expressed mdr1 and mdr3, the isoforms responsible for multidrug resistance. TKGM-F12 cells coexpressed mdr1 and mdr3 whereas TKGM-G2 cells expressed only mdr1. The drug transport function, measured by rhodamine 123 (R-123) efflux, was smaller in TKGM-F12 than in TKGM-G2 cells. The PGP substrates adriamycin, cyclosporin A, vinblastine, and verapamil inhibited R-123 transport in TKGM and TKGM-G2 cells. In the cells studied, PGP conferred some resistance to adriamycin; concomitant exposure to adriamycin with another PGP substrate impaired cell growth. The differential expression of mdr1 and mdr3 in mouse mesangial cell clones, the ability of mdr1 PGP to transport R-123, and the impairment of PGP-mediated transport in TKGM-F12 cells, coexpressing mdr1 and mdr3 products, are demonstrated. PGP may play a physiological role in mesangial cells.

Expression and function of P-glycoprotein in a mouse kidney cell line.

P-glycoprotein (PGP), a transporter conferring multidrug resistance to cancer cells, is expressed in the kidney. C219 monoclonal antibody binding revealed PGP in proximal tubules and mesangium of mouse kidneys. A cell line (TKPTS) expressing PGP was developed from proximal tubules of the 8Tg(SV40E)Bri7 mouse. Northern blot analysis demonstrated a 5.0-kb message identified as mdr1 by ribonuclease protection assay. Cyclosporin A (CSA) at 0.15 and 10 microM increased cellular accumulation of verapamil (VRP) by 32 and 121%, respectively (P < 0.001). VRP at 5 microM increased steady-state cellular accumulation of CSA by 46% (P = 0.02). Basal-to-apical transport of the PGP substrate vinblastine was inhibited by VRP. Rhodamine-123 (R-123) influx was rapid and independent of PGP. R-123 efflux was inhibited by VRP and CSA. Inhibition of PGP transport by VRP, CSA, and PSC-833 decreased the 50% effective dose of adriamycin. The concomitant administration of VRP and CSA was not deleterious and coincided with preferential accumulation of VRP over CSA. Inhibition of PGP-mediated transport is demonstrated as a mechanism of renal cell toxicity.