Neoexpression of ABH and Lewis blood group antigens in human hepatocellular carcinomas.

Expression of blood group ABH, Lewis, and sialylated-Lea antigens in human hepatocellular carcinomas and the adjacent nontumorous liver tissues was investigated with the use of seven monoclonal antibodies against these carbohydrate determinants. Chromatogram antibody-binding assay and solid-phase enzyme immunoassay of the upper-phase neutral glycolipids revealed the tumor-associated expression of blood group A-active glycolipids incompatible with blood-type status of the patients, a blood group A-active glycolipid with mobility on thin-layer chromatography between the known 6- and 8-sugar blood group A-active glycolipids in human erythrocytes, blood group H-active glycolipids, and blocked synthesis of Lea-active glycolipids with or without concomitant accumulation of Leb-active glycolipids. Immunohistochemical analysis of the fixed tissues with the use of an avidin-biotin-peroxidase complex method revealed blood group antigens in biliary epithelial cells but not in parenchymal liver cells. However, hepatocellular carcinoma cells in some cases expressed H and Leb antigens. Although only type 1 chain H antigen was detected in biliary epithelial cells, both type 1 and type 2 chain H antigens were found in hepatocellular carcinoma cells.

Neutralizing antibodies to human interleukin 6 reverse hypercalcemia associated with a human squamous carcinoma.

Bone resorbing cytokines may be associated with abnormalities in bone cell function and calcium homeostasis in a number of pathological conditions. One of these cytokines is interleukin 6 (IL-6), which is a multifunctional cytokine which has been shown to be associated with increased formation of bone resorbing osteoclasts in vitro. In this report, we demonstrate that neutralizing antibodies to human IL-6 lower the blood calcium in nude mice carrying a human tumor associated with increased IL-6 production. There was no effect on blood calcium in hypercalcemic nude mice carrying tumors not associated with increased IL-6 production and normocalcemic tumor-bearing nude mice. These results suggest that increased production of IL-6 may contribute to disturbances in calcium homeostasis in some malignancies, and suggest that neutralization of IL-6 effects can lower the serum calcium in these tumors.

A highly sensitive enzyme-linked immunosorbent assay for the measurement of interleukin-8 in biological fluids.

Interleukin-8 (IL-8) is a chemotactic and activating cytokine for neutrophils, which plays an important role in acute inflammatory responses. We aimed to develop a sensitive enzyme-linked immunosorbent assay (ELISA) for IL-8 and established 18 clones of anti-IL-8 monoclonal antibodies (mAbs). These mAbs were evaluated in terms of their antigen-binding affinities, and five clones were selected and used for the comparative study of various combinations of antibodies in sandwich ELISA. Affinity purified rabbit polyclonal antibody was also used in this study. One antibody pair, which showed relatively high sensitivity and which was not severely interfered with blood components, was selected and the assay conditions were optimized by choosing the appropriate buffer for sample dilution and by directly labeling the second antibody with enzyme. The finalized ELISA, using polyclonal antibody as first (coated) antibody and horseradish peroxidase-labeled mAb (clone EL139) Fab' fragment as second antibody, could detect as low as 2.5 pg/ml (0.125 pg/well) of IL-8 by in total 2 h incubation, without being affected by body fluid components. The ELISA was specific to IL-8, showing no cross-reactivity with other cytokines or various IL-8 family proteins which share some amino acid sequence homology with IL-8. As an example of its application to clinical specimens, plasma samples from patients with septic shock were measured. The results showed that sepsis patients contain significantly higher levels of plasma IL-8 compared to normal controls. When analyzed by gel-filtration chromatography, IL-8 in sepsis plasma was eluted in a molecular weight (M(r) region corresponding to the monomer form. The ELISA established here is expected to be effectively used for further investigations on the relationship between IL-8 and various diseases.

An enzyme-linked immunosorbent assay for the measurement of human interleukin-6.

An enzyme-linked immunosorbent assay has been developed to measure human interleukin-6. The assay, based on the avidin-biotin amplified two-step sandwich method, is quick (requiring 4.5 h), sensitive (detecting 9.5 pg/ml) and satisfactory in reproducibility and specificity. It shows good correspondence with the results of bioassays, and it is not affected by serum and plasma components. These results indicate that this ELISA is suitable for application to clinical samples, which is a major advantage over the widely used bioassays.

Ocular inserts for controlled release of antibiotics.

Ocular inserts impregnated with antibiotics (erythromycin and erythromycin estolate) which have sustained release characteristics were prepared, mainly for the purpose of trachoma therapy. In vitro experiments showed that the elution rate of a drug with low solubility in water (erythromycin estolate) is constant when the water content of the hydrogel insert is more than 30%. In the case of a drug with higher solubility (erythromycin), the elution rate depends on the water content. Some in vivo experiments using rabbit eyes were also reported.

Pseudomyxoma peritonei of ovarian origin--an autopsy case.

The autopsy case of an 80-year-old female with pseudomyxoma peritonei arising in the left ovary is reported. The patient was admitted with complaints of anorexia, sense of fullness and abdominal distension of two months' duration, and died of intestinal obstruction four months later. The autopsy revealed extensive tumor dissemination over the entire peritoneal cavity without any visceral invasion or distant metastasis. A part of the sigmoid colon showed marked stenosis and perforation with abscess formation. Histologically, the tumor was composed of various sized multiple cysts which were completely or incompletely lined by a layer of mucin-producing columnar epithelial cells with moderate nuclear hyperchromatism, and of a papillary pattern in some parts, indicating low grade malignancy.

Molecular cloning of the immunodominant regions of hepatitis C virus type III and IV by immunoscreening.

We have identified the immunodominant regions of hepatitis C virus (HCV) type III and IV by immunoscreening of a lambda gt11 cDNA library, which was constructed with RNA extracted from pooled sera of patients. In addition to the nucleocapsid protein, nonstructural region 3 (NS3), and NS4 regions, we found that the region at the N terminus of NS5 in type III and IV were also immunoreactive as well as in the prototype and type II, although amino acid homologies were approximately 60% between the prototype (or HCV type II) and HCV isolates of type III or IV. This result indicated that the region at the N terminus of NS5 may be a candidate for serotyping studies of HCV. This report presents the primary structure of immunodominant portions at the N terminus of NS5 in two HCV subtypes and a preliminary study of the immune responses to these antigens.

[A case of facial nerve neurinoma originated from the cerebellopontine angle portion].

Facial nerve neurinomas are relatively rare and most of them appear at the vertical portion of the facial nerve. Facial nerve neurinoma originated from the cerebellopontine (c-p) angle portion is less frequently reported. A 51-year-old woman was admitted to our hospital complaining of severe headache and nausea. She had had dizziness and unsteady gait for the previous two weeks. She did not complain of hearing disturbance, but otological examination revealed sensorineural deafness. She had no facial palsy. Skull x-ray showed no erosion of the internal auditory canal. Plain CT-scan revealed a large, unenhanced, low-density mass in the right c-p angle cistern. At the time of the operation, this tumor originated from the right facial nerve. Histological diagnosis of this tumor was schwannoma. After the operation, right facial nerve palsy appeared but hardness of hearing was no worse than previously. This tumor seemed to be facial nerve neurinoma in the c-p angle cistern. To the present, 121 facial nerve neurinomas have been reported in the previous literature. Facial nerve neurinomas in the c-p angle cistern, however, have only been reported in 5 cases. The most frequent symptom of facial nerve neurinoma in the temporal bone is facial nerve palsy, but that of facial nerve neurinoma in the c-p angle cistern is hearing loss, as in an acoustic neurinoma. Preoperative diagnosis of facial neurinoma in the c-p angle cistern using neurological symptoms alone is difficult. Furthermore, differential diagnosis from acoustic neurinoma in the c-p angle cistern using only skull x-rays and CT-scanning is also difficult.(ABSTRACT TRUNCATED AT 250 WORDS)

Phosphate in dialysis patients.

A single pool extracellular compartment (10-15 L) model was demonstrated to fit changes in plasma Pi (at steady state or rebound during HD) on the assumption that its G increased linearly or exponentially during HD. G at the end of HD was estimated to be 30-300 times that at the start of HD, which suggested rapid outflow of Pi from its reservoirs. When bovine or human RBC as a cellular model of probable Pi reservoirs were incubated with solute-free NSS or dialyzed plasma, BUN instantaneously reached a new equilibrium, but Pi and CR gradually flowed out from RBC. Pi efflux from RBC was observed even after the Pi concentration in the incubation media exceeded its initial plasma concentration. With regard to BUN and Cr, the concentration in RBC estimated from that in the incubation media decreased exponentially during HD corresponding to the change in their plasma concentrations. Estimated concentration of Pi in RBC, however, remained unchanged during HD in spite of significant decreases in its plasma concentration. These results suggest that Pi does not leave RBC by simple diffusion and that RBC contains Pi precursor(s). 31P NMR spectra obtained from packed RBC showed that uremic RBC contained more ATP and unidentified compounds (possibly sugar phosphates) than nonuremic RBC and that the peak amplitude of the latter unidentified compounds significantly decreased in uremic RBC leaving the dialyzer or at the end of HD. Furthermore, it was demonstrated that during incubation at 37 degrees C with NSS or dialyzed plasma, the peak amplitude of 2, 3-DPG decreased in contrast with an increased Pi peak.(ABSTRACT TRUNCATED AT 250 WORDS)

Dynamic behavior of plasma phosphate in chronic dialysis patients.

1. Change in the plasma P concentration during intra- and interdialytic phases is notably different from that for BUN, CR and UA. Reduction rate for P depends on its pretreatment concentration. Plasma P is apt to level off or rebound even during a treatment and quickly returns to the pretreatment level after it is terminated. 2. In short-term evaluations, occurrence of P rebound during a treatment does not correlate with factors such as meal, A1 gel, dialyzer type, dialyzer membrane and therapeutic mode, but with the pretreatment P concentration. Once it reaches a threshold level inherent to each patient, plasma P seems to rebound. 3. Pretreatment P concentration in each patient seemed to be controlled in a relatively narrow range. 4. While apparent generation rates (G) estimated with a single compartmental kinetic model are stable during intra- and interdialytic phases as to BUN, CR and UA, G for P seems to be significantly enhanced especially during the latter period of the treatment and immediately after the termination of the treatment.

Long-term observation on renal-osteodystrophy-related parameters in dialysis patients.

Among chronic haemodialysis patients, renal osteodystrophy is one of the most serious long-term complications which should be solved for substantially improving the quality of their lives. However, due to its multifactorial mechanism and the difficulty of its diagnosis, a final protocol to ameliorate and/or prevent renal osteodystrophy is difficult to establish. To discover any clues to a solution, we performed long-term observations on renal osteodystrophy-related parameters in chronic haemodialysis patients over 9 years, starting in 1980, focusing mainly on X-ray photodensitometry. Taking into consideration changes of treatments prescribed and/or tried during this period, changes of renal osteodystrophy-related parameters including metacarpal index, bone mineral content, and bone index were periodically monitored. While bone mineral content and bone index deteriorated for patients who were commenced on haemodialysis in 1980, during the consecutive 4 years, bone mineral content was significantly improved, with maintained bone index during the last 4 years for patients started on haemodialysis in 1985. Thirty-one male and thirteen female patients, monitored throughout the 9 years, showed a significant increase in bone mineral content during the past 4 years. The combined effects of current modality such as bicarbonate dialysate, active vitamin D3 and removal of accumulated aluminium, are suggested explanations.

High Rates of controlled ultrafiltration combined with optimal diffusion: recent advances in hemodialysis technique.

A system which permits high rates of middle molecule removal while preserving the removal of low molecular weight substances is described. It consists of a high fluid flux hollow fiber artificial kidney and an ultrafiltration controller. When used in combination, high rates of convective transport can be achieved through rapid ultrafiltration and reconstitution of blood with physiologic salt solutions. The theoretical impact of changing from conventional dialysis to this form of therapy on the body burden of middle molecules is estimated.

Rapid assay for detection of toxic shock syndrome toxin 1 from human sera.

A noncompetitive enzyme-linked immunosorbent assay that enables the quantitation of toxic shock syndrome toxin 1 (TSST-1) to as little as 30 pg/ml and the detection of TSST-1 to 10 pg/ml in phosphate-buffered saline including 33% human serum or plasma was developed. It takes only 3 h to complete this assay after plate preparation. In this study, 64 human serum samples obtained from 30 patients with toxic shock syndrome or toxic shock syndrome-like symptoms were subjected to testing for the detection of TSST-1. With a cutoff level for TSST-1 of less than 100 pg/ml, 28 samples obtained from 12 patients were positive for TSST-1. The mean and maximum concentrations for these TSST-1-positive samples were 440 and 5,450 pg/ml, respectively. Of these 12 patients, 8 were Staphylococcus aureus culture positive, 3 were negative upon bacterial culturing, and 1 had no cultures done.

Development of a new antithrombogenic continuous ultrafiltration system (ACUS) and its clinical evaluation.

A new totally antithrombogenic continuous ultrafiltration system (ACUS), consisting of polyacrylonitrile-polyethyleneoxide (PAN-PEO) membrane and ionically heparin-bound catheter, tubing, and module header was designed and its performance was confirmed through animal experiments and clinical evaluation. Animal experiments revealed that persistent antithrombogenicity and maintained ultrafiltration without systemic heparinisation were observed only when the three major parts--i.e. (1) the catheter and tubing, (2) the header part of a haemofilter module, and (3) the fibre membrane--were all antithrombogenic and connected to each other without uneven structures. ACUS was clinically applied to 15 oliguric patients with various severe conditions. We found that one filter could function for approximately 26 h without systemic anticoagulation, even in the presence of low blood pressure, and that ACUS did not change platelet function. Thus, ACUS seems to be very suitable for the management of volume overload, especially in patients with severe circulatory problems or bleeding tendencies.

Development of a new antithrombogenic continuous ultrafiltration system.

By interfacing a polyacrylonitrile (PAN)-polyethyleneoxide (PEO) membrane with an ionically heparin-bound catheter, tubing, and module header, a totally antithrombogenic continuous ultrafiltration system (ACUS) was developed and its performance, persistent antithrombogenicity, and well-maintained ultrafiltration level were confirmed through animal experiments. Although the amount of heparin released and accumulated in vitro from those heparinized parts was very low and stable (on the order of 1 x 10(-2) U/cm2/min), partial thromboplastin time evaluated in vivo was not elongated during passage through the ACUS. Extracorporeal circulation time with the ACUS in unheparinized dog model was 458 +/- 302 min (n = 24), whereas those of partially modified (antithrombogenic) system did not exceed 100 min. As compared with that in a conventional continuous arteriovenous hemofiltration system, an extracorporeal circulation with the ACUS in an unheparinized dog model revealed significantly less fluctuation of platelet count, and no adherent platelets were observed on the surface of the PAN-PEO membrane. An ACUS consisting of a PAN-PEO membrane and heparinized parts was thus demonstrated to have good platelet compatibility. An ACUS with a surface area of 0.25 m2 was applied to two patients with acute renal failure. Hemofiltration without systemic heparinization lasted for 44 h per hemofilter, and a stable level of ultrafiltration was maintained. This system seems to be applicable for the clinical management of volume overload, especially in patients with bleeding tendencies or postoperative bleeding.

Inhibitory effects of beta 2-microglobulin on in vitro calcification of osteoblastic cells.

Elevated levels of aluminum and beta 2-microglobulin have been demonstrated in chronic dialysis patients. The role of aluminum in the pathogenesis of renal osteodystrophy has also been shown. We report on the effects of beta 2-microglobulin on calcification in vitro using osteoblastic cells, clone MC3T3-El. At concentrations comparable to those in plasma of chronic dialysis patients, both beta 2-microglobulin and aluminum suppressed calcification while collagen synthesis and alkaline phosphatase activity were maintained. These observations may be related to the impaired bone mineralization frequently observed in chronic dialysis patients.

Removal of aluminum from chronic dialysis patients by administration of desferrioxamine and dialysis.

In order to perform quantitative in vitro and clinical studies on the removal of Al by the combined therapy of DFO administration and HD, an HPLC system for measuring DFO was established and a leaky membrane with enhanced permeability to middle molecular substances was developed. The DFO infusion caused rises in plasma Al levels (regarded as indicating the elution of accumulated Al) in patients undergoing HD. Plasma levels increased most in long-term patients and those with bone pain. Examination of Al clearances demonstrated that this increased plasma Al content passed through HD membranes at levels of more than 80 micrograms/l, and that the leaky membrane was more effective for removal of Al as well as DFO than the conventional one. Although a 1:1 complex between DFO and Al in an aqueous solution was confirmed, the formation of Al-rich complexes in vivo was suggested. It is concluded that Al-DFO complex formed in vivo can be effectively removed across the leaky membrane.

Globin chain synthesis in hemolytic anemia reticulocytes. A case of hemoglobin Burke.

Peripheral red blood cells from an anemic patient were incubated with [14C]Leu and the labeled globin chains were analyzed by CM-cellulose column chromatography in 8M urea. The radioactivity almost equal to that of beta chain emerged between beta and alpha chains. The newly appeared materials were found to be derived from an abnormal hemoglobin (Hb), Hb Burke [beta 107(G9)Gly to Arg], by the sequence analyses of abnormal globin chain and beta globin mRNA from the patient.

Clinical significance of a dialysis membrane that can remove beta 2-microglobulin (beta 2m).

Through clinical and in vitro evaluations clinical significance of continued use of a beta 2m removal membrane was validated.