[Further development of regional early care-Many roads lead to Rome : Developmental stages of four established rheumatological early care concepts in different regions of Germany]. / Regionale Frühversorgung weiterentwickeln ­ Viele Wege führen nach Rom : Entwicklungsetappen von 4 etablierten rheumatologischen Frühversorgungskonzepten in verschiedenen Regionen Deutschlands.

In order to shorten the prognostically relevant waiting time until diagnosis and initiation of appropriate treatment in inflammatory rheumatic diseases, rheumatological centers in many regions across Germany have established and continuously developed specific early care concepts. Evaluated models from Altötting·Burghausen, Berlin Buch, Düsseldorf and Heidelberg and their developmental stages as a response to internal and external challenges are presented in this overview. The transparent publication of the developmental steps and the exchange of experiences aim at promoting new early care concepts in other regions and continuing the joint dialogue for improvement of the early detection and quality of care of inflammatory rheumatic diseases in Germany.

[Early recognition and screening consultation: a necessary way to improve early detection and treatment in rheumatology? : Overview of the early recognition and screening consultation models for rheumatic and musculoskeletal diseases in Germany]. / Früh- und Screeningsprechstunden: Ein notwendiger Weg zur besseren Frühversorgung in der internistischen Rheumatologie? : Rheumatologische Früh- und Screeningsprechstundenmodelle in Deutschland.

In order to reduce the prognostically relevant time interval between the initial manifestation of a rheumatic and musculoskeletal disease and diagnosis as well as the consecutive initiation of an appropriate treatment, several rheumatological centers in Germany have improved the access to initial rheumatologic evaluation by establishing early recognition/screening clinics at their respective sites. Corresponding models located at Altoetting·Burghausen, Bad Pyrmont, Berlin Buch, Duesseldorf, Heidelberg, Herne, Mannheim as well as supraregional/multicenter initiatives Rheuma Rapid, RhePort and Rheuma-VOR are presented in this overview along with the respective characteristics, potential advantages and disadvantages, but also first evaluation results of several models. The aim of this publication is to promote early detection of rheumatic and musculoskeletal diseases as one of the most important challenges in current rheumatology by encouraging further rheumatologic centers and practices to launch their own early recognition/screening consultation model on the basis of aspects presented herein.

Action of DNA repair endonuclease ERCC1/XPF in living cells.

To study the nuclear organization and dynamics of nucleotide excision repair (NER), the endonuclease ERCC1/XPF (for excision repair cross complementation group 1/xeroderma pigmentosum group F) was tagged with green fluorescent protein and its mobility was monitored in living Chinese hamster ovary cells. In the absence of DNA damage, the complex moved freely through the nucleus, with a diffusion coefficient (15 +/- 5 square micrometers per second) consistent with its molecular size. Ultraviolet light-induced DNA damage caused a transient dose-dependent immobilization of ERCC1/XPF, likely due to engagement of the complex in a single repair event. After 4 minutes, the complex regained mobility. These results suggest (i) that NER operates by assembly of individual NER factors at sites of DNA damage rather than by preassembly of holocomplexes and (ii) that ERCC1/XPF participates in repair of DNA damage in a distributive fashion rather than by processive scanning of large genome segments.

Disruption of mouse ERCC1 results in a novel repair syndrome with growth failure, nuclear abnormalities and senescence.

BACKGROUND: The structure-specific ERCC1/XPF endonuclease complex that contains the ERCC1 and XPF subunits is implicated in the repair of two distinct types of lesions in DNA: nucleotide excision repair (NER) for ultraviolet-induced lesions and bulky chemical adducts; and recombination repair of the very genotoxic interstrand cross-links. RESULTS: Here, we present a detailed analysis of two types of mice with mutations in ERCC1, one in which the gene is 'knocked out', and one in which the encoded protein contains a seven amino-acid carboxy-terminal truncation. In addition to the previously reported symptoms of severe runting, abnormalities of liver nuclei and greatly reduced lifespan (which appeared less severe in the truncation mutant), both types of ERCC1-mutant mouse exhibited an absence of subcutaneous fat, early onset of ferritin deposition in the spleen, kidney malfunction, gross abnormalities of ploidy and cytoplasmic invaginations in nuclei of liver and kidney, and compromised NER and cross-link repair. We also found that heterozygosity for ERCC1 mutations did not appear to provide a selective advantage for chemically induced tumorigenesis. An important clue to the cause of the very severe ERCC1-mutant phenotypes is our finding that ERCC1-mutant cells undergo premature replicative senescence, unlike cells from mice with a defect only in NER. CONCLUSIONS: Our results strongly suggest that the accumulation in ERCC1-mutant mice of endogenously generated DNA interstrand cross-links, which are normally repaired by ERCC1-dependent recombination repair, underlies both the early onset of cell cycle arrest and polyploidy in the liver and kidney. Thus, our work provides an insight into the molecular basis of ageing and highlights the role of ERCC1 and interstrand DNA cross-links.

Effects of 24R,25-dihydroxyvitamin D3 in combination with 1 alpha-hydroxyvitamin D3 in predialysis renal insufficiency: biochemistry and histomorphometry of cancellous bone.

The effect of combined administration of 24R,25-dihydroxyvitamin D3 (24,25-(OH)2D3) and 1 alpha-hydroxyvitamin D3 (1 alpha-(OH)D3) was studied in 24 non-dialyzed patients with chronic renal insufficiency (CRI), matched pairwise as to age, sex, and creatinine clearance (Cr.cl). Low Ca intake had been supplemented beforehand. Then, 1 alpha-(OH)D3 (mean dose 0.55 micrograms daily) was given orally to all patients for 3 months (T0 to T3). Subsequently, patients were assigned randomly to 6 months further treatment either with 1 alpha-(OH)D3 alone (Group A) or with 1 alpha-(OH)D3 plus a high dosage of 24,25-(OH)2D3 (50 micrograms orally, twice weekly) (Group B). Histomorphometry was performed at T0, T3, and T9. In both groups iPTH was equally suppressed, into the lower normal range. Whereas in Group A, serum Ca rose steadily and Cr.cl declined, in Group B both parameters levelled off between T6 and T9. At T9, in Group A the elevated resorption and osteoid indices had normalized markedly, but osteoblasts (Ob.Pm) and mineralizing boundaries (M.Bd) were depressed considerably between T3 and T9. In contrast, in Group B, preservation of Ob.Pm and improved mineralizing activity were observed (M.Bd at T9 > T3 > T0). Resorption indices hardly changed. In the patients with high Ob.Pm at T0, cancellous bone area increased significantly. This was not observed in Group A. Thus, in Group B, osteoblast recruitment appeared maintained and M.Bd appeared normalized. Decline of remodeling toward an adynamic state with an increased risk of hypercalcemia appeared prevented.

Changes of interstitial bone thickness with age in men and women.

Mean interstitial bone thickness has been estimated from mean trabecular thickness and mean wall thickness (It.Th = Tb.Th-2*W.Th) with varying results. Some authors found age-related changes of It.Th while others did not. We measured It. Width (It.Wi, 2D) directly, in cancellous iliac crest bone from 23 women aged 20-78 yrs. and 21 men aged 23-74 yrs. At grid-selected sites coupled measurements were done of Tb.Wi and It.Wi, together with W.Wi. It.Wi was defined as the distance between cement lines nearest to the surface at both sides of a trabecula. A thionine stain was used to visualize the cement lines. When the persons studied were divided into two age groups: 20-49 yrs (Group 1) and 50-79 yrs (Group 2) significant but opposite age-related trends were seen both in men and women. Group 1: Men: r = -.68; p < .02; n = 11; Women: r = -.68; p < .02; n = 11. Group 2: Men: r = .79; p < .01; n = 10. Women: r = .59; p < .05; n = 12 (r = coeff. of correlation). Men: It.Wi 63.3 microns (3rd decade [dec.]); 50.8 microns (6th dec.); 67.8 microns (8th dec.). Women: It.Wi 63.7 microns (3rd dec.); 48.3 microns (6th dec.); 63.8 microns (8th dec.). Measured It.Wi values appeared considerably larger than their calculated counterparts (mean delta: 35%). The decline of It.Wi is interpreted as a sign of negative bone balance at the BMU level, leading to thinning of trabeculae, the subsequent increase has to be the result of both declining resorption depth and disappearance of the thinner trabeculae.(ABSTRACT TRUNCATED AT 250 WORDS)

Immunolocalization and quantification of noncollagenous bone matrix proteins in methylmethacrylate-embedded adult human bone in combination with histomorphometry.

The noncollagenous proteins (NCPs) in the bone matrix comprise growth factors with distinct cellular effects and a series of proteins with less clear biological actions. In order to understand the role of these proteins in bone metabolism and in bone diseases, it is crucial to determine their localization and quantity in normal and pathological bone. We have developed an immunohistochemical method to detect osteopontin, osteocalcin, bone sialoprotein, osteonectin, decorin, biglycan, and the growth factors transforming growth factor-beta, insulin-like growth factor-I, and bone morphogenetic protein-2 both in bone matrix and in bone cells of adult human bone embedded in methylmethacrylate. Immunohistochemistry and standard bone histomorphometry in adjacent sections allows the localization of the proteins to metabolically active sites in bone. The protocol works with several fixatives and with bone specimens obtained and embedded to over 20 years ago. Most importantly, we developed a procedure to specifically stain the mineralized matrix green in combination with a red staining of the NCPs. Using digital image analysis it is possible to quantify the relative amounts of NCPs (microm2 NCP area/microm2 mineralized matrix area). Within one biopsy of normal bone cut at four different heights (at a distance of 100 microm), two adjacent sections were stained either for osteopontin or osteonectin. Thirty trabecular and 20 cortical microscopic fields were measured, and the NCP:mineralized matrix ratio was calculated. Stepwise analysis of the standard error of the mean of the NCP:mineralized matrix ratios showed that measuring about 50 microscopic fields is sufficient to obtain representative data with a small confidence interval. In conclusion, the present procedure enables to quantify NCPs and to relate their presence to metabolically active sites in bone. The quantification provides the opportunity to monitor differences in distribution (e.g., cortical vs. trabecular) and differences between normal and pathological conditions and to assess changes in matrix composition during treatment. This can be done by reanalyzing bone biopsies obtained in the past, e.g., during clinical trials. Therefore, the present technique will be a valuable tool for the study of noncollagenous bone matrix proteins in human bone.

Role of macrophages in nephrolithiasis in rats: an analysis of the renal interstitium.

Interstitial calcium oxalate (CaOx) crystals can be found in primary oxalosis and in secondary hyperoxaluria. In a rat model for nephrolithiasis, we investigated whether such crystals can be removed by the surrounding interstitial cells. CaOx crystals were induced by a crystal-inducing diet based on ethylene glycol (EG) and ammonium chloride (CID). Both lithogenic compounds were added to the drinking water. After 9 days, the animals received normal drinking water for 2 days. Using this CID, only the interstitial crystals are retained. Subsequently, half of the population remained on normal drinking water (normo-oxaluria), whereas the other half received a low dose of EG alone (chronic hyperoxaluria). The rats were killed at regular times thereafter. The results showed that the kidney-associated oxalate significantly declined during normo-oxaluria, but remained high during chronic hyperoxaluria. Interstitial cells positive for the leukocyte common antigen (CD45; which identifies all types of leukocytes), the ED1 antigen (which is specific for monocytes and macrophages), and the major histocompatibility class II antigen (MCHII), respectively, had increased in number, with minor differences between both rat populations. The cells around the interstitial crystals were mostly positive for ED1. Multinucleate giant cells were regularly observed. These cells were positive for CD45 and ED1 and sometimes also for MCHII. The crystals in these cells were moderately positive for acid phosphatase and carbonic anhydrase II. It is concluded that interstitial CaOx crystals can be removed under normo-oxaluric conditions and that, in all likelihood, macrophages and multinucleate giant cells are involved in that process.

Cytokine production induced by binding and processing of calcium oxalate crystals in cultured macrophages.

Deposition of calcium oxalate (CaOx) crystals in the renal interstitium is common in humans with primary oxalosis and secondary hyperoxaluria, as well as in kidneys of rats with CaOx nephrolithiasis. In vivo, macrophages and multinucleated giant cells mostly encapsulate these crystals. To investigate whether macrophages are able to dispose of CaOx crystals after phagocytosis, we used a nontransformed macrophage cell line derived from mouse spleen progenitors. Cytokine assays showed that in response to crystal binding and phagocytosis, these macrophages release tumor necrosis factor-alpha. This release was evident at 8 hours, maximal at 24 hours, and decreased to control values after 48 hours of incubation with crystals. A very low but significant release of interleukin-6 into the culture medium was only noticed after 32 hours. Radiochemical experiments showed that these cells bind 38.8% of the CaOx crystals added. After 4 days, all internalized crystals had been dissolved and their molecular constituents released into the extracellular environment. Confocal laser scanning microscopy followed by morphometrical analyses confirmed these results. Long-term (survival) analyses showed that in the interval under study and at the crystal doses used, cell viability was not significantly affected. These findings support the view that properly functioning macrophages are able to remove CaOx deposits from the renal interstitium and that these cells produce inflammatory cytokines before crystal dissolution.

Quantitative analysis of the decay of immunoreactivity in stored prostate needle biopsy sections.

Application of immunohistochemistry to assess the presence of prognostic tissue markers is used widely. The quantitation of these markers may be hampered by a time-related loss of antigenicity in formalin-fixed paraffin-embedded tissue stored on glass slides. Potential loss of immunohistochemical staining intensity was studied on prostatic needle biopsy sections stored for a maximum of 4 years with antibodies against p27kip1, CD-44s, MIB-1, and androgen receptor (AR). In benign tissue, the positive/total ratio for p27kip1 was determined, while CD-44s staining intensity was assessed semiquantitatively. For MIB-1 and AR, nuclear staining intensity was assessed using computed image analysis. An exponential and significant decay of immunoreactivity was seen for p27kip1, CD-44s, MIB-1, and AR, with half-lives of 587 days, 214 days, and 290 days for p27kip1, MIB-1, and AR, respectively. Immunohistochemical assessment of prognostic tissue markers on stored slides must be considered with care in research and clinical settings.

Treatment of relapsing Mycobacterium avium infection with interferon-gamma and interleukin-2 in an HIV-negative patient with low CD4 syndrome.

A patient with idiopathic CD4 T-lymphopenia was diagnosed with a recurrent disseminated Mycobacterium avium infection. Because of progressive disease, treatment with interferon-gamma (IFN-γ) and interleukin-2 (IL-2) was started. The patient was successfully treated with IFN-γ-1b and IL-2 in addition to anti-mycobacterial combination therapy. To our knowledge, this is the first report of successful combination therapy with IFN-γ-1b and IL-2 in a patient with idiopathic CD4 T-lymphopenia. Short-term IFN-γ-1b and IL-2 might be considered as therapeutic options in refractory mycobacterial infections in patients with idiopathic CD4 lymphopenia.

[Recurring disseminated Mycobacterium avium infections in an HIV-negative patient]. / Rezidivierende disseminierte Mycobacterium-avium-Infektionen bei einem HIV-negativen Patienten.

HISTORY AND ADMISSION FINDINGS: A 64 year old man complained about progressive pain in his right upper leg and pelvis since 4 weeks without any history of trauma. He noticed a reduced general health status for several months with weight loss and subfebrile temperatures. The patient's medical history revealed several malignant tumours and recurrent infections with atypical mycobacteria and herpes viruses. INVESTIGATIONS: Laboratory findings showed signs of chronic inflammation. Multiple disseminated osteolytic bone lesions, hepatosplenomegaly, lymphadenopathy and multiple splenic lesions were detected by CT scan. Mycobacterium avium was isolated from an osteolytic lesion of the pelvic bone. Malignant disease and acquired immunodeficiency syndrome were excluded. Lymphocyte counts showed lymphocytopenia with reduced T cells, B cells and a reduced CD4-/CD8-ratio. Interferon-gamma-pathway defects as described in patients with susceptibility to atypical mycobacteria could not be identified. TREATMENT AND COURSE: Clinical and immunologic findings indicated a link between recurrent Mycobacterium avium infections and an idiopathic CD-4 T-cell lymphopenia (low CD4 syndrome). Antimycobacterial treatment at standard doses was started according to the resistance test obtained from microbiological culture. Medical and radiological checkup after 3 months of therapy showed signs of progressive disease. CONCLUSIONS: Osteolytic lesions can be caused by mycobacterium avium infection. Disseminated atypical mycobacteriosis is an opportunistic disease in patients with congenital or acquired immunodeficiency syndrome.

Quantitative immunohistochemistry of androgen receptors in a microsphere model system and in prostate tissue sections.

OBJECTIVE: To evaluate androgen receptor (AR) levels to predict endocrine therapy response and prognosis. STUDY DESIGN: A sepharose microsphere model was employed to establish the relationship between level of immunohistochemical staining density and both antigen and primary antibody (F39.4.1) concentration. Subsequently, the results of the model system were compared with the results in routine prostate sections. RESULTS: A log-linear relationship was observed between optical density (OD) and primary antibody dilution measured in immunostained, antigen-coated microspheres with moderate antigen density. Similar titration curves were observed in prostate sections at the same dilution range, indicating that the microsphere model can be extrapolated to routine tissue sections. With microspheres with high coating density, the loglinear range of dilutions shifted to higher dilutions. CONCLUSION: Differences in AR levels in prostate tissue sections might be more accurately detected by comparison of titration curves of primary antibody than by comparison of OD values at a fixed primary antibody dilution.