The presently described study was undertaken to modify our solid-phase, two-site enzyme immunoassay method (EIA) for human pancreatic secretory trypsin inhibitor (PSTI) to make it more practical for the quantification of the inhibitor in human body fluids. Peroxidase activity fixed on the solid phase was measured spectrophotometrically with 2, 2′-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) as a coupling reagent. The analytical range of the assay was from 4 to 1, 000ng/ml when a sample volume of 50μl was used, and PSTI levels so measured in serum correlated well with those measured by conventional radioimmunoassay (n=16, r=0.985, intercept +10.35, and slope 1.12). Both the anti-PSTI monoclonal antibody IgG-coated polystyrene beads and the anti-PSTI monoclonal antibody Fab'-linked peroxidase conjugates were found to be stable for at least 4 months at 4°C. Our evaluation of the resulting method revealed that the procedure is acceptable in regard to linearity, recovery, precision, and specificity. In addition to these qualities the EIA method for PSTI introduced here is reliable and rapid enough to be useful for either routine determinations and/or clinical tests.
Two forms of prolidase could be separated by TSK DEAE-5PW chromatography from erythrocytes of normal subjects and from those of the mother of two patients with prolidase deficiency. Only one form of prolidase was separated from erythrocytes of the patients with prolidase deficiency. Each peak (I and II) of prolidase differed in its response to Mn2+, substrate specificity, and heat stability. Prolidase activity in erythrocytes from the patients showed a complete lack of peak I of normal prolidase, whereas peak II had normal activity against all the substrates tested. Molecular weights of partially purified prolidases I and II from normal erythrocytes were about 110, 000 and 185, 000 daltons, respectively and that of the prolidase from erythrocytes of the patients, 185, 000 daltons. The various properties between patients' prolidase and peak II of normal prolidase were very similar. The activity of peak I from erythrocytes of the patients' mother was reduced about 50 compared with that of normal individuals, but activity of peak II was almost the same. These results suggest that for at least the present two cases, the lower prolidase activity seen in erythrocytes from patients with prolidase deficiency results from lack of the peak I enzyme rather than from an alteration of normal prolidase.
The nutritional status of 30 Egyptian sprayers occupationally exposed to pesticides was assessed by a variety of blood chemistries and a number of other tests and measurements. The results showed that younger workers with a mean age of 14 years had poorer overall nutritional status with significantly lower (p<0.05) mean hemoglobin, plasma phospholipids, and retinol-binding protein concentrations compared with the corresponding mean values obtained for sprayers with a mean age of 33 years. Plasma retinol level correlated significantly (p<0.05) with age, body weight, plasma lipids, cholesterol, phospholipids, alanine amino transferase activity, and retinol-binding protein. Further, frequent exposure of the sprayers to pesticides led to a significant increase in the mean blood urea level, and to a decrease in the mean levels of plasma lipid and carotene.
Blood selenium concentrations were found to be significantly lower in patients with chronic pancreatitis than in normal controls. The activity of the selenoenzyme erythrocyte glutathione peroxidase had a significant positive correlation with selenium concentration and it was also significantly lower in patients with the disease. These findings suggest that selenium deficiency and reduced glutathione peroxidase activity are associated with the elevation of lipid peroxides in chronic pancreatitis.
The cause of enormous accumulation of arachidonic acid (AA) in the epidermis of psoriasis (a benign epidermal hyperproliferative disease) patients is unknown. As an initial step to elucidate the reason, the fatty acid composition in whole plasma lipid extract of psoriatic and normal subjects, male and female separately, was studied by gas chromatography. The results revealed a decrease in plasma linoleic acid (LA), a major component of whole plasma fatty acid, in both male and female psoriatic patients as the only statistically significant finding. A statistically significant decrease in AA in psoriatic plasma as reported previously was not confirmed in males. The results suggest that plasma LA in psoriatic patients is selectively incorporated into epidermis and transformed into AA there, so as to meet the great demand of psoriatic epidermis for AA. However, another possibility is also considered that a decrease in LA itself is the cause of psoriasis, since animals fed an essential fatty acid-deficient diet show psoriasis-like skin and their epidermal DNA synthesis is increased. Interestingly, increased tendency of plasma docosahexaenoic acid (DCHA) level in psoriatic patients was also found, which has not been reported before. Just as indomethacin exacerbates psoriasis, DCHA, which was shown to be a strong competitive inhibitor of AA conversion to prostaglandins, may be involved in the development of psoriasis.
The effect of serum obtained from Arthus reaction-elicited rabbits on the generation of reactive oxygen species (ROS) by normal rabbits' polymorphonuclear leukocytes (PMNs) was examined. The oxidative metabolism of infiltrating PMNs in the skin during the Arthus reaction was observed by the nitroblue tetrazolium dye reduction test (NBT test). The significantly increased H2O2 generation by normal PMNs was observed when PMNs were incubated with serum obtained 24 and 48h after the elicitation. Also the increased O2- generation was noted after incubation with serum taken at 48h after the elicitation. Many NBT-positive PMNs were observed in skin specimens obtained 24 and 48h after the elicitation. These results may indicate the importance of ROS generated by infiltrating PMNs in immune complex vasculitis because the Arthus reaction is a model of immune complex vasculitis.
Levels of mouse epidermal growth factor (mEGF) in mouse plasma were measured by a sensitive two-site enzyme immunoassay (EIA) to study whether sialoadenectomy (removal of submaxillary gland) affects the level of the factor in the circulation. mEGF levels in plasma of sialoadenectomized mice were lower than those of sham-operated (operated without removal of submaxillary gland) mice, but the difference was not remarkable. That is, the submaxillary gland is not the sole source of EGF in the mouse body. Though the reproductive ability of normal male mice and of those sialoadenectomized or sham-operated was also studied, no significant difference in fertility rate of the mated female mice was found among them. Thus the EGF in submaxillary gland seems not to play important roles in male reproductive function of the mice.
Effects of vitamin E (VE) on tumor growth and in vitro lymphocyte proliferation as markers of cellular immunity in CDF1 mice were examined in the VE-deficient state as well as in the VE-sufficient state attained by pretreatment with VE. Oral (585mg of dl-α-tocopheryl nicotinate per 100g diet) or intraperitoneal (0.5ml of saline-diluted d-α-tocopherol every other day at a dose of 40mg/kg) supplementation with VE did not enhance in vitro lymphocyte proliferation to concanavalin A, phytohemagglutinin, or lipopolysaccharide, suppress the growth of Meth A fibrosarcoma, or prolong the survival of tumor-bearing animals, even though the serum VE value was maintained 1.5 to 2.4 times higher in the group fed the VE-sufficient diet and about 3 times higher in the group injected with VE than the value for the group fed the control diet. On the other hand, VE-deficiency produced by a basal VE-deficient diet (0.16mg of d-α-tocopherol per 100g diet) seemed to be disadvantageous for the tumor-bearing host in view of the decrease in in vitro lymphocyte proliferation to phytohemagglutinin or lipopolysaccharide, rapid tumor growth, and shortened survival of tumor-bearing mice.
This study evaluates the effect of continuous fluid infusion on metabolic alterations in septic rats. The saline administration, designed to maintain blood volume, was carried out from the onset of peritonitis induced by cecal ligation and puncture. The operated rats were divided into two groups: the physiological saline-resuscitated group, treated by continuous infusion intravenously (2.0ml/h) immediately after surgery, and the untreated septic group. Concentrations of hepatic adenine nucleotides, glycogen, glucose-6-phosphate, phosphoenolpyruvate, lactate, malate, RNA, DNA, and protein, were determined at the times of 4, 7, 12, and 24h. Alterations in free amino acid levels in plasma and liver were also measured. A rapid decrease in plasma glucogenic amino acids and perturbation of carbohydrate metabolism were noted at 4h and alterations in metabolite levels became evident at 7h in both septic rat groups, when compared with those in normal rats. In addition, obvious differences in the alteration patterns of metabolite contents were observed at 12h between the two septic rat groups. The untreated septic rats appeared to be in a shock state at 12h, showing approximately 50% mortality, whereas all septic rats with resuscitation still survived at this time. These results indicate that an adverse effect of lowered tissue perfusion was involved in further aggravation of hepatic ATP-producing metabolism in the untreated septic rats.
Lipid peroxide levels in the skin of the senescence-accelerated mouse (SAM) were examined in relation to the appearance of symptoms in the skin. In SAM-P/1 mice (accelerated senescence-prone mice), symptoms such as a decrease in hair glossiness and skin ulcers begin to be observable at 5-6 months of age, mainly in the skin of dorsal neck. Prior to the appearance of the skin symptoms, the lipid peroxide level in this region of the skin of SAM-P/1 mice was significantly higher than that of the corresponding region of SAM-R/1 mice (accelerated senescence-resistant mice). Lipid peroxide levels in the skin of dorsal neck tended to be in parallel with those in the serum in SAM-P/1 mice of 3-4 months of age. These results suggest that in SAM-P/1 mice lipid peroxides increased in the blood are transferred to the skin where they provoke degeneration of the cells followed by the appearance of skin lesions.