[Consequences of autopsies for the living : Causes of death in the clinical diagnosis "septic and toxic shock"]. / Konsequenzen von Autopsiebefunden für die Lebenden : Todesursachenfeststellungen bei der klinischen Diagnose "septisch-toxischer Schock".

BACKGROUND: There is reason to believe that the diagnosis of septic and toxic shock, as indicated on the death certificate, cannot be confirmed as the cause of death without autopsy and subsequent histological analysis. The external examination of the corpse can therefore not represent the sole basis for a reliable statement about the infection status of a corpse, e. g. as a prerequisite for embalming. MATERIAL AND METHODS: The validity of autopsy in determining septic and toxic shock as the cause of death is demonstrated in 7 exemplary cases. RESULTS: Decades of experience in a university pathology institute have shown that an external examination of the corpse alone is not suitable for certifying the cause of death if an infectious disease is suspected. Consequently, only autopsy with subsequent histological analysis provides reliable statements on the etiopathogenesis of the underlying process. Possible problems and discrepancies between clinical and pathological diagnoses are discussed on the basis of several cases with or without autoptic confirmation of the septic shock. The case of a missionary from Africa infected with Lassa virus serves to point out the seriousness of the threat an undiagnosed infection may represent to the attending staff. CONCLUSION: During the treatment of patients suspected to have an infectious cause of fever of unknown origin, compliance with the usual safety regulations, including adequate disinfecting measures, is essential. In cases with fatal outcome, not infrequently under the clinical picture of a septic and toxic shock, autopsy should be regularly performed to confirm the type of infection and the infectious cause of death. Rapid and open communication between the professional groups involved plays a crucial role in this process.

In vivo behavior of murine epidermal cell lines derived from initiated and noninitiated skin.

The in vivo behavior of cell cultures derived from normal and carcinogen-treated mouse epidermis was studied by implanting the cultures in a s.c. vascularized bed protected by a silicone chamber. Cells derived from normal adult mouse epidermis as well as cells derived from tumor-promoter-treated skin were unable to grow in these systems. Conversely, cell lines derived from skin initiated with single doses of N-methyl-N'-nitro-N-nitrosoguanidine or 9,10-dimethyl-1,2-benzanthracene proliferated in these chambers, reforming an epithelial structure. The type of structure in the chambers varied, ranging from formation of almost normal epithelia to atypical invasive behavior. The variable in vivo behavior among the different cell lines may be attributed to the initiation agent, the number of passages of the cultures, random genetic events, the strain of mouse, or a combination of these factors. Most of the cell types used in this study and all the cell lines that were able to grow in these chambers were selected for resistance to Ca-induced terminal differentiation. However, resistance to terminal differentiation according to the Ca2+ switch does not always correlate with the ability to grow in the chambers, since cell lines derived from spontaneous foci of resistance failed to grow in this system. These studies showed some of the possibilities of the SC silicone chambers to study the histogenic potential of cell lines derived from carcinogen-treated epidermis. This system also appears suitable to study the complex relationship between epidermal cells and specialized (dermal) stroma.

Enhanced malignant progression of mouse skin tumors by the free-radical generator benzoyl peroxide.

Chemical carcinogenesis in mouse skin can be divided into the processes of initiation, promotion, and progression. The free-radical generator benzoyl peroxide is moderately active during the promotion stage. Repetitive treatment of mouse benign skin tumors (papillomas) with benzoyl peroxide (20 mg, twice weekly) increased the number of cumulative carcinomas per group by 325% and the number of keratoacanthomas by 44% compared to tumor-bearing Sencar mice treated with the promoter 12-O-tetradecanoylphorbol-13-acetate. The lack of increase in the number of cumulative papillomas per group due to benzoyl peroxide treatment suggests that benzoyl peroxide enhanced the progression of preexisting papillomas. The ability of benzoyl peroxide to enhance the progression of benign tumors to cancer should be considered when determining the human risk from exposure to this widely used chemical agent; in addition, biological assays specifically testing malignant progression may be essential and beneficial for determining an agent's carcinogenic risk.

Nucleotide sequence of rat vascular cell adhesion molecule-1 cDNA.

Vascular cell adhesion molecule 1 (VCAM-1) is an inducible transmembrane protein which is expressed by vascular endothelium following cytokine activation. VCAM-1 mediated the adhesion of certain blood leukocytes and tumor cells via the interaction with its counter-receptor, the integrin VLA4. When initially cloned from interleukin-1 (IL-1) stimulated human umbilical vein endothelial cells, VCAM-1 was reported to contain six immunoglobulin-like domains. However, subsequent cDNA clones and structural analysis of the human gene evealed an alternatively spliced seventh immunoglobulin domain. This seven domain form appears to be the predominant transcript in IL-1 activated endothelium. In this report, the cloning and nucleotide sequence of rat VCAM-1 is described.

High-dose azathioprine pulse therapy as a new treatment option in patients with active Wegener's granulomatosis and lupus nephritis refractory or intolerant to cyclophosphamide.

The objective of this study was to evaluate the feasibility and safety of high-dose azathioprine pulse (HAP) therapy in the induction of remission in patients with active Wegener's granulomatosis (WG) or progressive lupus nephritis (LN) refractory to or intolerant of cyclophosphamide. Four patients with antineutrophil cytoplasmic antibody (ANCA)-associated WG and two patients with progressive LN were treated with HAP (1200-1800 mg) applied monthly as continuous intravenous infusions at 50 mg/h. Patients received a total of 50 courses of intravenous azathioprine (AZA) therapy. Disease activity was assessed using the Birmingham Vasculitis Activity Score (BVAS) and the Systemic Lupus Erythematosus Activity Index (SLEDAI). As only partial remission was induced in patients with progressive LN on this regimen, an additional 18 cycles were applied in these patients in which oral AZA at 100 mg/day in weeks 2 and 3 was added between two intravenous courses. A hereditary defect in thiopurine methyltransferase activity was excluded before initiation of treatment. High-dose azathioprine pulse and the intensified HAP treatment were well tolerated. Complete remission was achieved in two patients with WG suffering from three relapses of disease on application of 2-6 courses of HAP. Remission was maintained for 16-24 months. The remaining two patients with WG were withdrawn after 2-3 courses due to unchanged disease activity. In two patients with LN, partial remission was noted on 6-9 courses of HAP; however, the patients relapsed despite therapy with methotrexate and mycophenolate mofetil. The intensified HAP regimen led to partial or complete remission in both LN patients which was confirmed by sequential renal biopsies. Our results suggest that HAP therapy represents a well-tolerated regimen in patients with active WG and LN intolerant of or refractory to cyclophosphamide. As partial or complete remission was observed in four of six patients, further studies seem warranted to assess clinical efficacy in these patients.

Malignant progression of mouse skin papillomas treated with ethylnitrosourea, N-methyl-N'-nitro-N-nitrosoguanidine, or 12-O-tetradecanoylphorbol-13-acetate.

Mouse skin tumors were induced by a single topical application of 7,12-dimethylbenzanthracene (DMBA), followed by biweekly promotion with 12-O-tetradecanoylphorbol-13-acetate (TPA). After 20 weeks of promotion, mice were treated twice weekly for 2 weeks with either ethylnitrosourea (ENU), N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) or TPA. Thereafter all groups were treated biweekly with TPA. The ENU-treated group had a higher percentage of animals with carcinomas and developed 217% more cumulative carcinomas per group than TPA-treated controls. The percentage of mice with carcinomas and the cumulative number of carcinomas per group in MNNG-treated mice was higher than TPA-treated controls but was less than ENU-treated mice. The ratio of cumulative carcinomas to cumulative papillomas in ENU treated, MNNG-treated and TPA-treated mice was 16%, 9% and 6%, respectively. Histological examination of tumors remaining at the termination of the experiment revealed the presence of keratoacanthomas, some of which stained positive for gamma-glutamyltransferase (GGT), in the ENU-treated and MNNG-treated, but not the TPA-treated groups. The fact that no new papillomas developed during the progression stage indicated that enhanced carcinogenesis resulted from the progression of pre-existing tumors. Enhanced progression of benign skin tumors in mice by only a few treatments of an agent may serve as a potential model for studies into the mechanisms and the inhibition of malignant progression. The model also allows for a comparison of the potency of agents in enhancing malignant progression.

Allogeneic transplantation of normal epidermal cells and squamous cell carcinomas in SENCAR mice.

A method used in our laboratory for the transplantation of single cell suspensions of epidermis and skin carcinomas is described. Silicone chambers were placed into a subcutaneous granulation tissue formed by the implantation of a glass disk. The skin was closed on top of the chamber by using Michael autoclips. Cells were injected 24 to 48 hr later through the skin using a syringe and needle. The neoformation of epithelia with adnexa was observed when newborn epidermal cells were injected. When a fresh cell suspension of squamous cell carcinoma was used, a typical differentiated carcinoma was formed within 2 to 4 weeks. However, after 4 weeks, some of the grafts showed mild signs of rejection. The technique described is a useful system to transplant squamous cell carcinomas and can also be used as a rapid assay for malignancy.

ETS-dependent p16INK4a and p21waf1/cip1 gene expression upon endothelin-1 stimulation in malignant versus and non-malignant proximal tubule cells.

AIM: Cellular senescence, leading to cell death through prevention of regular cell renewal, is associated with the upregulation of the tumor suppressor gene p16(INK4a). While this mechanism has been described as leading to progressive nephron loss, p16(INK4a) upregulation in renal cell carcinoma has been linked to a disease-specific improved patient survival rate. While in both conditions endothelin-1 is also upregulated, the signaling pathway connecting ET-1 to p16(INK4a) has not been characterized until this study. MAIN METHODS: Cell culture, qRT-PCR, Western Blot, immunoprecipitation (IP), proximity ligation assay (PLA), and non-radioactive electrophoretic mobility shift assay (EMSA). KEY FINDINGS: In malignant renal proximal tumor cells (Caki-1), an activation of p16(INK4a) and p21(waf1/cip1) was observed. An increased expression of E-26 transformation-specific (ETS) transcription factors was detectable. Using specific antibodies, a complex formation between ETS1 and extracellular signal-regulated kinase-2 (ERK2) was shown. A further complex partner was Mxi2. EMSA with supershift analysis for ETS1 and Mxi2 indicated the involvement of both factors in the protein-DNA interaction. After specifically blocking the endothelin receptors, ETS1 expression was significantly downregulated. However, the endothelin B receptor dependent downregulation was stronger than that of the A receptor. In contrast, primary proximal tubule cells showed a nuclear decrease after ET-1 stimulation. This indicates that other ETS members may be involved in the observed p16(INK4a) upregulation (as described in the literature). SIGNIFICANCE: ETS1, ERK2 and Mxi2 are important complex partners initiating increased p16(INK4a) and p21w(af1/cip1) activation in renal tumor cells.

Recovery of whisking function after manual stimulation of denervated vibrissal muscles requires brain-derived neurotrophic factor and its receptor tyrosine kinase B.

Functional recovery following facial nerve injury is poor. Neuromuscular junctions (NMJs) are "bridged" by terminal Schwann cells and numerous regenerating axonal sprouts. We have shown that this poly-innervation of NMJs can be reduced by manual stimulation (MS) with restoration of whisking function. In addition, we have recently reported that insulin-like growth factor-1 (IGF-1) is required to mediate the beneficial effects of MS. Here we extend our findings to brain derived neurotrophic factor (BDNF). We then examined the effect of MS after facial-facial anastomosis (FFA) in heterozygous mice deficient in BDNF (BDNF(+/-)) or in its receptor TrkB (TrkB(+/-)). We quantified vibrissal motor performance and the percentage of NMJ bridged by S100-positive terminal Schwann cells. In intact BDNF(+/-) or TrkB(+/-) mice and their wild type (WT) littermates, there were no differences in vibrissal whisking nor in the percentage of bridged NMJ (0% in each genotype). After FFA and handling alone (i.e. no MS) in WT animals, vibrissal whisking amplitude was reduced (60% lower than intact) and the percentage of bridged NMJ increased (27% more than intact). MS improved both the amplitude of vibrissal whisking (not significantly different from intact) and the percentage of bridged NMJ (11% more than intact). After FFA and handling in BDNF(+/-) or TrkB(+/-) mice, whisking amplitude was again reduced (53% and 60% lower than intact) and proportion of bridged NMJ increased (24% and 29% more than intact). However, MS failed to improve outcome in both heterozygous strains (whisking amplitude 55% and 58% lower than intact; proportion of bridged NMJ 27% and 18% more than intact). We conclude that BDNF and TRkB are required to mediate the effects of MS on target muscle reinnervation and recovery of whisking function.

Recovery of whisking function promoted by manual stimulation of the vibrissal muscles after facial nerve injury requires insulin-like growth factor 1 (IGF-1).

Recently, we showed that manual stimulation (MS) of denervated vibrissal muscles enhanced functional recovery following facial nerve cut and suture (FFA) by reducing poly-innervation at the neuro-muscular junctions (NMJ). Although the cellular correlates of poly-innervation are established, with terminal Schwann cells (TSC) processes attracting axon sprouts to "bridge" adjacent NMJ, molecular correlates are poorly understood. Since quantitative RT-PCR revealed a rapid increase of IGF-1 mRNA in denervated muscles, we examined the effect of daily MS for 2 months after FFA in IGF-1(+/-) heterozygous mice; controls were wild-type (WT) littermates including intact animals. We quantified vibrissal motor performance and the percentage of NMJ bridged by S100-positive TSC. There were no differences between intact WT and IGF-1(+/-) mice for vibrissal whisking amplitude (48 degrees and 49 degrees ) or the percentage of bridged NMJ (0%). After FFA and handling alone (i.e. no MS) in WT animals, vibrissal whisking amplitude was reduced (60% lower than intact) and the percentage of bridged NMJ increased (42% more than intact). MS improved both the amplitude of vibrissal whisking (not significantly different from intact) and the percentage of bridged NMJ (12% more than intact). After FFA and handling in IGF-1(+/-) mice, the pattern was similar (whisking amplitude 57% lower than intact; proportion of bridged NMJ 42% more than intact). However, MS did not improve outcome (whisking amplitude 47% lower than intact; proportion of bridged NMJ 40% more than intact). We conclude that IGF-I is required to mediate the effects of MS on target muscle reinnervation and recovery of whisking function.

Hepatic steatosis in organ donors: disparity between surgery and histology?

BACKGROUND: In times of organ shortage, use of marginal cadaveric livers has become increasingly important to reduce pressing organ demand and rising death rates while awaiting donations. Indisputably, fatty change in donor livers is a risk factor for poor initial function after orthotopic transplantation. However, identifying and rejecting marginal from good donor livers is one of the most difficult surgical tasks. Unfortunately, a liver biopsy with rapid histological diagnosis is rarely performed to identify marginal livers. METHODS: From 2005 to 2008, we investigated 36 livers of organ donors, which were explanted but not transplanted or underwent liver wedge biopsy during organ donation. All livers underwent standard surgical procedures and were allocated by Eurotransplant International Foundation. After unsuccessful allocation, explanted livers were photographically documented, formalin-fixed, and analyzed histopathologically. RESULTS: Seven livers were classified as good organ quality by the surgeon (19.4%); 15 were acceptable (41.6%); and 14 poor (39%). In 63.8% of livers, a frozen section was performed; 6/36 cases (16.7%) showed macrovesicular and microvesicular steatosis of less than 30%. In addition, all six cases fulfilled two or less extended donor criteria, as defined by the German Medical Association. CONCLUSION: More marginal livers from cadaveric organ donors could have been transplanted. To extend the transplant pool of liver grafts, liver biopsies should be performed in all cases of acceptable and poor livers. If frozen section analysis is performed, a wedge liver biopsy should be taken from at least two different segments of the liver to validate the histological results.

Hypereosinophilic syndrome associated with acute necrotizing myocarditis and cardiomyopathy.

We report the rare case of a 55-year-old female with massive eosinophilic myocarditis and severe, however reversible, impairment of left ventricular function. The patient presented with reduced physical condition, progressive dyspnea on exertion and peripheral edema. The white blood count revealed a leukocytosis and markedly elevated peripheral blood eosinophilics (48.8%). An endomyocardial biopsy demonstrated massive myocardial infiltration with eosinophilic granulocytes and necrosis. The symptoms and laboratory parameters indicate the presence of a hypereosinophilic syndrome. The differential diagnosis of a Churg-Strauss syndrome is discussed. Medical heart failure treatment according to international guidelines and an immunosuppressive treatment with prednisolone (Decortin H) 1.5 mg/kgBW) were initiated. This therapy led to a dramatic reduction of the eosinophilic granulocyte count and normalization of the peripheral blood count, which correlated with a significant improvement of clinical symptoms. Consistently, an increase of left-ventricular function was observed. Upon successive dose reduction to a maintenance dosage of 10 mg prednisolone, the patient's clinical status and peripheral blood count remained stable.

Radiation therapy induced modulation of wound healing at experimental vein graft anastomoses.

OBJECTIVES: The aim of this study was to investigate if radiation therapy (RT) favorably modulates wound healing at vein graft anastomoses. MATERIALS AND METHODS: Jugular vein grafts were sewn into carotid arteries in 32 rats which were randomly divided into two groups: RT (gamma source, 14 Gray, n=16) and control (C, sham irradiation, n=16). Grafts and adjacent arteries were analyzed at 2 (n=8) and 8 weeks (n=8) by histology, immunohistochemistry, and morphometry. RESULTS: Although, RT did not reduce the overall occurrence of intimal hyperplasia, the distribution differed. RT led to a reduction of intimal hyperplasia in arterial segments (median: C: 41.873 microm2; RT: 6.452 microm2, p < 0.0007). In contrast, RT augmented intimal hyperplasia in vein grafts (median: C: 30.287 microm2; RT: 90.455 microm2, p < 0.014). Vein graft diameters after RT were enlarged (median: C: 2.098 microm; RT: 3.381, p < 0.031). Over 80% of the cells were of mesenchymal origin in both groups. CONCLUSIONS: RT reduced intimal hyperplasia in arterial segments. However, RT led to graft dilatation and increased intimal hyperplasia in vein grafts. RT did not favorably modulate the vascular wound healing response in this model.

Night radiology.

Night radiology is the practice of the in-hospital radiologist from 4:00 to 11:00 p.m. His duty is to keep the interpretation of radiographs current. At the medical center described, an average of 95 radiographic examinations per day are performed during the evening and at night; 60 of these require immediate interpretation. Night radiology was instituted because of the large number of unmonitored and uninterpreted films that had to be dealt with the following morning. The night radiology duty is seven consecutive nights and is rotated among all nine staff radiologists. Night radiology provides a service that the emergency department and the private physician can rely on and can use without hesitation, delay, or resistance.

Alterations of glomerular podocytic processes in immunologically mediated glomerular disorders.

The importance of the relationship between the alteration of podocytic processes (PP) and proteinuria has been controversial for almost 30 years. In spite of several morphometric studies the range of PP alterations in different proteinuric conditions has gone unnoticed. Thus, we studied the distribution of abnormally broadened PP in different glomerulopathies and in normal controls (children and adults). We found a highly variable mean PP-width in all proteinuric patients, values that were, however, always clearly distinct from normal controls in children and adults. A correlation between the extent of PP alteration and the amount of proteinuria was not found (r = 0.09) for the group as a whole or for the different clinico-pathological entities. In proteinuric patients 15 to 50% of all PP were broadened and covered 40 to 60% of the total length of the peripheral capillary loop. We conclude that the alteration of PP in various proteinuric conditions is variable and does not correlate with the level of proteinuria. These studies further suggest that other mechanisms, possible hemodynamic adaptation in response to the capillary wall injury, might play a role in the permeability changes observed in these patients.

Nuclear factor-kappa B mediates induction of vascular cell adhesion molecule-1 in glomerular mesangial cells.

Cultured glomerular mesangial cells (GMCs) can be activated at the transcriptional level by a variety of physiologically relevant factors including cytokines, endotoxin and glycosylated end products. The mechanism with which the signal is transduced from the membrane to the nucleus of these cells is largely unclear. In vascular endothelial cells, the signal transduction pathway involves activation of the pleuripotent transcription factor, NF-kappa B, and leads to increased expression of a variety of genes including vascular cell adhesion molecule-1 (VCAM-1). Here, we demonstrate that TNF-alpha and IL-1 beta transiently induced VCAM-1 mRNA expression in a time dependent manner. TNF-alpha also induced the specific interaction of proteins from GMC nuclei with an oligonucleotide bearing the NF-kappa B binding sites in the VCAM-1 promoter. Electrophoretic mobility shift and supershift analysis indicated that the p65 subunit of NF-kappa B is a component of this induced complex. Finally, reporter activity driven by a VCAM-1 promoter-chloramphenicol acetyltransferase reporter construct increased 8-10 fold following TNF-alpha incubation, or p65 cotransfection. Thus, the p65 subunit of NF-kappa B is activated in GMCs exposed to cytokine and can mediate induction of gene expression.

Determinants of immune complex-mediated glomerulonephritis.

We have studied the influence of steric factors on the clinico-pathologic expression of immune complex-mediated glomerular diseases, utilizing ferritin as an exogenous antigen. The tracer was planted in the left kidney either in the subepithelial layer of the glomerular capillary wall or on the endothelium and lamina rara interna. Subepithelial immune complex formation resulted in non-inflammatory injury with heterologous and autologous proteinuric phases (115 +/- 16 mg/24 hrs on day 2; 183 +/- 16 mg/24 hrs on day 9) lasting four to five weeks. The glomerular filtration rate of the experimental left kidney was reduced by 19% at day 3, and was increased by 20% at day 12 over right kidney values. Immune complexes persisted for more than seven weeks in the lamina rara externa. In contrast, immune complex deposition on the endothelium and in the lamina rara interna led to acute transient anuria, with a 38% drop in glomerular filtration rate at one hour, massive platelet accumulation, followed by a strong inflammatory response. Proteinuria did not develop. Functional and structural integrity was restored within 24 hours, with complete clearing of immune deposits. We conclude that the distribution of exogenous antigens within the capillary wall determines the structural and functional expression of immune-mediated glomerular diseases.