The chronic myelocytic cell line K 562 contains minor (m) as well as major (M) ber/abl fusion mRNAs.

By searching for additional chimeric bcr/abl transcripts in K 562 cells characterized by major (M) bcr/abl fusions, a new mRNA, a minor (m) bcr/abl transcript, was detected. A practical implication of this finding is that the K 562 cell line can be used as positive control for the detection by the polymerase chain reaction of both types of transcripts for the diagnosis of Philadelphia chromosome associated leukemias.

Quality assurance in RT-PCR-based BCR/ABL diagnostics--results of an interlaboratory test and a standardization approach.

Here we describe the results of an interlaboratory test for RT-PCR-based BCR/ABL analysis. The test was organized in two parts. The number of participating laboratories in the first and second part was 27 and 20, respectively. In the first part samples containing various concentrations of plasmids with the ela2, b2a2 or b3a2 BCR/ABL transcripts were analyzed by PCR. In the second part of the test, cell samples containing various concentrations of BCR/ABL-positive cells were analyzed by RT-PCR. Overall PCR sensitivity was sufficient in approximately 90% of the tests, but a significant number of false positive results were obtained. There were significant differences in sensitivity in the cell-based analysis between the various participants. The results are discussed, and proposals are made regarding the choice of primers, controls, conditions for RNA extraction and reverse transcription.

Clinical relevance of hepatitis G virus (HGV) infection in heart transplant patients.

To investigate whether the recently discovered hepatitis G virus (HGV) influences the clinical outcome of heart transplant recipients under immunosuppression, we determined the prevalence of HGV infections correlated with liver function and survival in 51 patients. Presence of HGV RNA and anti-E2, a marker for resolved HGV infection, were serially tested in sera from patients before and after heart transplantation (HTX) by nested RT-PCR and ELISA. Four of 51 (7.8%) patients before transplantation, and 22 of 50 patients (44%) after transplantation showed signs of persistent or resolved HGV infection. HGV infection was not associated with impairment of liver function or with patient survival. In summary, presence of HGV infection does not influence the clinical outcome in heart transplant patients.

Survival distribution in breast cancer.

It has been stated that breast cancer survival rates follow an exponential distribution. This would mean that the mortality rate is constant. Survival distribution was analyzed by the clinical life table method in one series of 10,752 patients and in another of 656 patients followed up to 8 and 18 years, respectively. Part of the larger series' table is (table: see text). Necessarily, clinical survival data are censored progressively. These kinds of data are analyzed best by examining the hazard function, which is the instantaneous death rate, or force of mortality. If an exponential distribution described survival in breast cancer correctly, the hazard function would be constant. These data clearly are not consistent with an exponential distribution, as the hazard function decreases. The survival distribution calculated from these data shows that the chance of dying of cancer decreases the longer a patient survives. This is more optimistic and consistent with clinical experience than is the exponential distribution.

Prostacyclin (epoprostenol): its effect on canine splanchnic blood flow during hemorrhagic shock.

Prostacyclin (epoprostenol, prostaglandin I2) is a vasodilator of the splanchnic circulation during normotensive states. To confirm the persistence of its effects after hemorrhagic shock, six anesthetized, previously splenectomized, adult mongrel dogs were subjected to hemorrhagic shock using a modified Wigger's technique in which a mean arterial pressure of 30 mm Hg was maintained until 25% of the shed blood spontaneously returned. The animals were randomly resuscitated with normal saline solution or a similar volume of saline solution containing prostacyclin. Organ blood flow was calculated by measuring the distribution of radioactively tagged microspheres. During shock, blood flows to the liver, small intestine, pancreas, and carcass were reduced. During a 60-minute infusion, prostacyclin selectively caused a significant increase in hepatic arterial blood flow. This improvement in arterial blood flow may prove beneficial in the clinical management of hemorrhagic shock.

Virus-binding activity of fibronectin: masking of hepatitis A virus.

Human plasma fibronectin interacts with viruses. When fibronectin-containing human sera negative for antibodies to hepatitis A virus (HAV) were added to suspensions of HAV, radioimmunological detection of HAV was reduced. This masking effect seemed to depend on the fibronectin concentration of the sera: plasma fibronectin purified by cryoprecipitation and affinity chromatography showed a masking effect on purified HAV which was dependent on the concentrations of fibronectin and HAV. Fibronectin peptides were obtained by subtilisin digestion: the non-collagen-binding regions of the fibronectin molecule were involved in the binding of HAV. We conclude that fibronectin has a virus-binding activity which interferes with radioimmunological methods for virus detection, and may contribute to the frequent transmission of hepatitis viruses by blood products enriched in fibronectin.

Cutaneous papulo-vesicular eruptions in non-A, non-B hepatitis.

A relapsing papulo-vesicular rash with or without pruritus was observed in 54 out of 148 patients (36%) with posttransfusion or sporadic, acute or chronic, non-A, non-B (NANB) hepatitis. The predominant location was the trunk and the anterior surfaces of the upper extremities. The face was affected less often. The eruptive phase was accompanied by general symptoms and increases in aminotransferases and gamma-GT values. The nature of the eruption was consistent with cutaneous reactions as frequently seen in enterovirus infections. No predominance was found for special groups of patients when the skin lesions were correlated to either sex, mode of infection or pattern of transaminase elevation (i.e. monophasic or bi-, and multi-phasic).

Parameters predicting response to alpha-interferon treatment in chronic hepatitis C.

BACKGROUND/AIMS: Sustained response to alpha-interferon treatment for chronic hepatitis C is seen in only 25% of cases. Therefore, it is desirable to define pretreatment factors predicting responders. MATERIALS AND METHODS: Forty-nine patients with chronic hepatitis C were treated with a standard alpha-interferon regimen (3 x 3 MU s.c./week). Demographic, biochemical and immunological parameters, and HCV genotypes were obtained prior to initiation of treatment and evaluated for their value in predicting response to alpha-interferon therapy. RESULTS: Response, as defined by normalization of ALT, was 71% during interferon therapy and sustained response after discontinuation of interferon 24.5%. Patients infected with HCV-genotype 1b had significantly more often "community-acquired" disease. Their outcome was worse with a response rate of 44% during therapy and a sustained response of 12.5%, as compared to 87% and 27% respectively in patients infected with genotypes other than 1b. On multivariate analysis, absence of cirrhosis, HCV-genotype other than 1b, higher ALT levels and higher numbers of CD8 positive liver infiltrates were found to be predictors of response during alpha-interferon therapy. CONCLUSION: Response to alpha-interferon therapy seems to be influenced both by viral virulence factors and by the intensity of the host immune response to HCV.

Vertebral erosions in patients undergoing maintenance hemodialysis for chronic renal failure.

Unlike articular erosions in the appendicular skeleton, the frequency, appearance, and clinical relevance of vertebral erosions in azotemic osteodystrophy are not known. Lateral vertebral radiographs of 118 patients on maintenance hemodialysis were reviewed to assess the frequency, distribution, and rate of progression of vertebral erosions. Thirty (25%) of 118 patients showed a superficial corner erosion resembling the so-called Romanus lesion of ankylosing spondylitis. Their presence, whether at a single level or at multiple levels, did not correlate with erosive changes in the phalanges or sacroiliac joints that are known to occur in renal osteodystrophy. Vertebral erosions may progress slowly with time, but seem to have limited bearing on clinical symptoms. In two patients, however, the radiographic changes progressed dramatically, simulating an infection. Biopsy and surgical intervention were considered but not carried out because one patient declined and in the other an infected graft was identified as the source of infection. Both patients had negative scintiscans, and their subsequent clinical courses excluded infectious spondylitis. Vertebral erosions in patients on maintenance hemodialysis are frequent, and there appears to be a spectrum of changes from superficial erosions to large resorptive defects. Further diagnostic evaluation of these erosive changes appears to be warranted only rarely.

Complement-mediated acinar cell necroses in pancreatitis induced by basement membrane antibodies.

To study the role of the serum complement system in the early necrosis of acinar cells an acute pancreatitis was produced by injection of basement membrane antibodies into the pancreatic duct of mice and rats. In all animals deposition of complement (C3) and antibasement membrane IgG could be observed in an identical position within areas of acinar cell necrosis. The extent of parenchymal damage and the intensity of complement deposits corresponded to the injected dose of antibodies. The importance of cytolytically active complement components (C5-9) was demonstrated in congenitally C5-defective old line mice which did not show typical centrolobular necroses 1 hr after intraductal injection of antibodies. However, the normocomplementemic mice developed extensive necroses of acinar cells. The results support the hypothesis of a complement-induced acinar cell necrosis in acute pancreatitis.

[Rational use of serological methods in diagnosis of virus hepatitis A and B (author's transl)]. / Rationeller Einsatz serologischer Methoden zur Untersuchung der Virushepatitis A und B.

The serological diagnosis of virus hepatitis A and B depends on the demonstration of the various virus antigens (HA-antigen, HBs-antigen, HBe-antigen) and on their corresponding antibodies (HA-, IgG- and IgM-antibody, HBc-antibody, Hbe-antibody and HBs-antibody). The knowledge of epidemiology of hepatitis and characteristic fluctuation of hepatitis-markers in the course of the disease allows not only statements on the aetiology, prognosis and the phase of hepatitis but also may help to avoid unnecessary expensive investigations and loss of time in diagnosis.

[Immunohistological investigations and complement analysis in immunological diseases (author's transl)]. / Immunhistologische Untersuchungen und Komplementanalysen bei immunologischen Erkrankungen.

Immunohistological techniques and complement analysis are essential methods of investigation in the diagnosis of immunologically induced disease processes today. Immunopathogenesis or participation of immunological reactions in inflammatory processes such as glomerulonephritis, dermatitis, arthritis for example, can be detected through immunohistological demonstration of deposits of immunoglobulin and complement in biopsy material. Numerous antibodies against tissue and infection pathogens can be demonstrated in the serum by indirect immunofluorescence techniques.

Virus inactivation and prevalence of GBV-C in haemophiliacs.

A new putative hepatitis virus has recently been discovered and termed GB virus C (GBV-C). We investigated the prevalence of this virus among 50 haemophiliacs treated with non-virus-inactivated clotting factor concentrates prior to 1985 and 21 haemophiliacs treated exclusively with virus-inactivated clotting factor concentrates. In the first group the prevalence of GBV-C based on PCR and ELISA was 46%. In the second group the prevalence of GBV-C was similar to that of healthy blood donors (5%). We therefore conclude that GBV-C is reliably inactivated by modern virucidal methods such as vapour heating, pasteurization and treatment with solvent/detergent mixtures.

[Hepatitis non-A, non-B. Retro- and prospective studies on the epidemiology of the acute disease]. / Hepatitis non-A, non-B. Retro- und prospektive Untersuchungen zur Epidemiologie der akuten Erkrankung.

The epidemiology was studied in 159 consecutively admitted patients (1981-1983) with acute and chronic parenteral and non-parenteral type non A, non B hepatitis (HNANB). To establish the frequencies of types A (HAV), B (HBV) and HNANB data were collected from the official health statistic of the Federal Republic of Germany (1980-1982). Accordingly, 5 out of 100 000 persons acquired HNANB each year. There was no regional prevalence (i. e. industrial areas, cities) in HNANB as it was present in HAV. The relation of HNANB to HAV and HBV was 1:3:2,4 (health statistic), but was 1:1,1:3 in clinical studies (1979-1983, n = 2027). Of the patients with non-parenteral HNANB (n = 50) most were elder than 20 years of age. Susceptibility for parenteral acquired HNANB was observed in all groups of age. The evaluation of the possible modes of infection revealed 59% of the patients with non-parenteral ("sporadic") type HNANB, other 28,9% had posttransfusion HNANB. When monthly incidence was examined the HNANB infectivity of bloodtransfusions was high during May, June and October. Most cases of sporadic HNANB became clinically ill during summer and October. Difficulties in determination of incubation periods became evident because of the fluctuating increases and decreases of aminotransferases: In posttransfusion HNANB (n = 28) the first increases of aminotransferases were recorded at day 18 +/- 13 whereas peak values happened at day 77 +/- 21. It is concluded, that HNANB is an infection which affects a high proportion of the population, and needs consideration as a common infectious disease.

[Endotoxinemia in acute pancreatitis (author's transl)]. / Endotoxinämie bei akuter Pankreatitis.

In a clinical study the question was asked, whether endotoxinemia of intestinal origin accompanying acute pancreatitis would influence decisively the course of this disease. In 21 of 32 patients with acute pancreatitis endotoxinemia could be diagnosed by the Limulus test, in this group 9 patients died. In the group of patients without endotoxinemia however no patient died. Extrapancreatic complications as for instance coagulation defects, renal failure, and pulmonary changes were predominant in the patients with endotoxinemia. From these findings the conclusion is made, that endotoxinemia occurs frequently in acute pancreatitis and influences decisively the course of the disease.

The cyclic behaviour of NANB hepatitis as basis for standardized diagnostic.

Efforts were made to characterize the clinical and biochemical behaviour of NANB hepatitis in 51 patients. 15 patients had posttransfusion NANB hepatitis, 36 a sporadic form of the disease. The patients' complaints predominantly were nausea and vomiting (64%), in about each 25% cardial complaints, lassitude, muscle pain and fever were observed. An eczema in a kind of maculopapular eruptions was frequently seen. The ratio of SGOT/SGPT was almost never less 1.0, Gamma-GT was consistently increased. The biochemical changes relapsed frequently. In posttransfusion NANB hepatitis the relapses were observed to occur predominantly around day 21, 28, 42, 49, 56, 63, 70, 77, and in further weekly intervals.

[Hepatitis C virus: sequence homology of a European isolate and divergence from the prototype]. / Hepatitis-C-Virus: Sequenzhomologie europäischer Isolate und Divergenz zum Prototyp.

The polymerase chain reaction (PCR) detected specific hepatitis C viral (HCV) RNA sequences in liver biopsies from two patients with chronic hepatitis, in the tissue of a liver implantate, in plasma from four chronic non-A, non-B hepatitis (NANBH) patients and, for the first time, in an infectious anti-D-immunoglobulin preparation. A comparison of the viral sequences coding for a region for the nonstructural NS3 protein from the liver tissues revealed only a very small degree of sequence divergence on the cDNA as well as on the amino acid level (between 0 and 5%). The sequence similarities of the RNA isolated from plasma of the four chronic NANBH patients and the anti-D-immunoglobulin preparation were partly somewhat lower but altogether also high (between 90 and 100%). In contrast, all eight cDNA and amino acid sequences exhibited a significantly higher degree of divergence in comparison with the HCV prototype sequence (between 29 and 32%) than among themselves (between 0 and 10%). This unexpected high sequence similarity of the eight European isolates and their low homology to the Northamerican prototype sequence is indicative for the existence of different types of HCV. This will be important not only for epidemiological studies but also for the development of effective diagnostic procedures and vaccines. Concerning the pathogenesis of NANBH, a double infection or a helper mechanism has to be considered: in addition to the C virus, sequences of an other virus particle were found in the infectious IgG preparation as well as in the liver biopsies.

Hepatitis C virus infections in dialysis units: prevalence of HCV-RNA and antibodies to HCV.

HCV infection causes serious complications in dialysis patients that lead to problems in management of patients in dialysis units. Determination of HCV-RNA is at present essential for monitoring the course of HCV infection. Reports concerning HCV-RNA in dialysis patients are mostly from Asian dialysis units; therefore, an analysis of dialysis patients in Europe was undertaken. From 1515 patients 2630 blood samples were screened for HCV-RNA and anti-HCV. Two-thirds of patients positive in an anti-HCV test containing a mixture of three antigens (EIA-II, Ortho) were further analysed for antibodies against these individual antigens. From 523 patients multiple samples were tested. Related on dialysis units from which all the attending patients were tested, mean prevalence of HCV-RNA was 8.4%, of anti-HCV 13.2%. Concerning all plasma samples from dialysis patients sent to our laboratory for investigation of HCV-RNA and anti-HCV the prevalence of HCV-RNA was 21.9%, of anti-HCV 23.1%, HCV-RNA was present in 76% of anti-HCV positive patients and in 4.1% of anti-HCV negative patients (1.3% of single and 6.8% of multiple tested patients). Acute and chronic infections with self-limited, persistent or intermittent viraemia were observed with changes and fluctuations of both HCV markers. With the exception of differences in onset of antibody production in some patients following acute infections, there were no major differences of dialysis patients compared to patients without dialysis treatment as far as antibody spectrum and detectability of HCV-RNA were concerned.

Antibodies reacting with the glomerular mesangium. Isolation and immunopathology.

Antigens of porcine lung were prepared by homogenization and ultrasonic treatment of lung tissue. The antigens were digested by collagenase and trypsin and chromatographed on Sephadex G 200. After immunization of rabbits with certain fractions of the chromatographed material, antibodies could be obtained in rabbits which fixed in vitro and in vivo in the glomerular mesangium of rats, hogs and the human, but not in the basement membrane of glomeruli of these species. Following intravenous injection into rats, the antibodies could be observed in the mesangial area without an apparent loss of antigenicity for 55 days. While 5 days after the application of 20 mg of the anti-mesangial IgG-preparation, there were no glomerular changes histologically demonstrable, after 55 days small amounts of rat-IgG could be demonstrated histochemically within the mesangium. Deposits of rat-complement (C3) could not be demonstrated with certainty. Severe morphological lesions were absent. Rats injected with the same doses of antibody and simultaneously immunized with rabbit-IgG showed a substantially greater deposit of autologous rat-IgG and rat-complement within the mesangial area after 55 days. By histological examination focal and segmental scleroses of the mesangium were determined. A significant pathological proteinuria did not occur. The present model constitutes a new possibility for studying the function of the glomerular mesangium.