In vitro diagnosis of penicillin and streptomycin allergy. The specific leukocyte alteration reaction using luminescence microscopy]

The method of fluorescent microscopy was used for studying the diagnostic value of the reaction of the leucocyte specific alteration (LSA) in patients with different syndromes of hypersensitivity, allergy in the anamnesis and without hypersensitivity to penicillin and streptomycin. It was found that only markedly positive results of the LSA reaction (independent of the sensibilization type) were of diagnostic value, the results of the reaction being stated in half of the patient with hypersensitivity in the anamnesis and in 3/5 of the patients with allergy. Simultaneous use of other tube immunological or skin tests was recommended for the other patients with lower levels of the positive results of the LSA reaction with a purpose of etiological diagnostics or revealing latent sensibilization before treatment with the antibiotics. The LSA reaction is recommended for practical use in complex with other methods of allergological examination.     

Reversing the detrimental effect of adriamycin on skin grafts in rats

We evaluated in a rat model the effects of a homologous fibrin glue in reversing the effects of Adriamycin on adherence and take of skin grafts. A total of 40 male Fisher rats were used in the study. During the first phase of the experiment, the animals were assigned to either group I (N = 10) receiving normal saline or group II (N = 10) receiving 6 mg/kg Adriamycin by tail vein injection 24 hours before surgery. Skin grafts with and without fibrin glue were placed over wounds in the backs of the animals and adherence was measured at 24 and 48 hours. In the second phase (N = 20), the experiment was repeated, this time evaluating the total area of skin graft take at 7 days. Fibrin glue was found to increase adherence and take of skin grafts in all Adriamycin-treated animals.     

Upregulation of glucose metabolism by granulocyte-monocyte colony-stimulating factor

Alterations of glucose metabolism were investigated for 6 hours following an intraarterial injection of murine recombinant granulocyte-monocyte colony-stimulating factor (GM-CSF) (30 micrograms/kg body weight). GM-CSF resulted in a transient elevation of plasma glucose. The rate of whole body glucose appearance, as measured by infusion of [6-3H] glucose, was increased by about 10% between 0.5 and 3 hours following GM-CSF injection. In vivo glucose utilization of individual tissues was investigated by the tracer 2-deoxyglucose technique. At 30 min, GM-CSF increased glucose utilization by 80-90% in liver and lung, and 50-60% in skin and spleen. At 3 and 6 hours, glucose utilization by these tissues returned toward control levels except for lung. There was a 40-50% increase in glucose utilization by skeletal muscle 30 min after GM-CSF which was sustained for 6 hours. Glucose utilization of testis, ileum and kidney did not change significantly. Plasma concentrations of insulin, glucagon and tumor necrosis factor were not altered in response to GM-CSF. These findings indicate that some of the acute metabolic effects of a short-term administration of GM-CSF are observed in macrophage-rich tissues, and suggest that GM-CSF may be involved in the metabolic upregulation of immunologically active tissues.     

DNA replication in mammalian cells exposed to the action of physical, chemical or biological factors. IV. 5-fluorodeoxyuridine modifies the effect of hyperthermia on DNA synthesis and the survival of HeLa cells

Preliminary incubation of logarithmically growing HeLa cells with FUdR decreases an inhibitory effect of hyperthermia (43 degrees C, 1 hour) on DNA synthesis. The hyperthermia alone inhibits DNA synthesis considerably: the label in acid-precipitable material accounts for 30% of control level. Preliminary incubation of the cells with FUdR (10(-6)) for 24 or 6 hours (plus 18 hours in fresh medium) decreases the effect: the label yields account for 50 or 90% of the respective control levels. A molecular weight of nascent DNA synthetized in the cells after hyperthermia or incubation with FUdR is lower than the control one but it increases rapidly during postincubation. Nucleoid of cells treated with FUdR has a sedimentation velocity which exceeds that of the control cells by more than 25%. Preliminary incubation with FUdR sensitizes the cells to hyperthermia. The effect is not believed to be associated with cells synchronization since the treatment of the cells with FUdR for 2 or 6 hours, when FUdR itself does not exert its toxic effect, brings about sensibilization of cells to hyperthermia. It is suggested that modification of the cell viability and DNA replication are related to some changes of chromatine structure induced by FUdR.     

The state of lipid peroxidation and of the antioxidant system in the liver and lung microsomes at different periods of sensitization depending on the histamine content]

Histamine impact on lipid peroxidation (LPO) in hepatic and pulmonary microsomes of guinea-pigs sensibilization was studied. It is demonstrated that intensity of chemiluminescence and LPO product content in NADPH-, ascorbat-dependent and spontaneous systems was enhanced in relation to the period of sensibilization process formation and histamine concentration.     

Toxic effects of catecholamines on skin

The purpose of this study was to examine the effects of catecholamines on skin necrosis independent of their vasoactive effects. Rat abdominal or human breast skin was excised, pinned flat, and incubated at 37 degrees C for 6 hours in a buffered salt solution containing catecholamine. At 0.1 and 6 hours the lactate dehydrogenase (LDH) released from the skin and appearing in the buffer was determined spectrophotometrically. All groups showed similar LDH levels at 0.1 hour. Rat skin treated with greater than or equal to 10(-7) M epinephrine (33 times less than the 1:200,000 used clinically) or greater than or equal to 10(-5) M norepinephrine showed a significant increase in the LDH released at 6 hours versus controls (18.75 +/- 1.25 versus 13.75 +/- 1.25 and 29.25 +/- 2.96 versus 22.00 +/- 1.96 IV, respectively). Total tissue LDH levels were not significantly different at 0.1 or 6 hours. The toxic effect of epinephrine was eliminated by the addition of propranolol or selective beta 2 blockade, but not by alpha or beta 1 blockade. Therefore, this effect appears to be mediated largely by beta 2 receptors. Similar toxic effects were seen in human breast skin treated with 1:200,000 epinephrine and were blocked with propranolol. Phenylephrine at 1:20,000 demonstrated toxicity, but angiotensin II and vasopressin did not. These studies indicate that addition of catecholamine to ischemic rat or human skin accelerates skin death within 6 hours, but that the toxicity can be reversed with beta blockade.     

Intradermal injection of Hsp60 induces cytokine responses in canine atopic and healthy skin

The purpose of this study was to investigate the immunoregulatory potential of Hsp60 in the skin of dogs with atopic dermatitis. Three dogs with chronic atopic dermatitis and four healthy dogs were injected intradermally with Hsp60 and phosphate-buffered saline. Biopsies were taken before testing from non-injected control skin, lesional and non-lesional atopic skin, and 48 and 72 h after injection. Analysis of cytokine messenger RNA was performed using quantitative real-time polymerase chain reaction. Forty-eight hours after Hsp60 injection, a rise in interleukin (IL)-10 was found (P = 0.034) with the highest expression levels in non-lesional atopic and control skin. A rise of transforming growth factor beta (P = 0.015) and IL-12p40 (P = 0.017) was noticed 72 h after Hsp60 injection in control skin. No significant differences were observed for the expression of IL-4, IL-12p35, and interferon gamma. The results indicate that Hsp60 is able to induce cytokines of a regulatory and Th1 phenotype in the skin. Furthermore, this study seems to provide a first indication of deficient Hsp60 response in atopic dermatitis affected skin.     

Fate of interleukin-6 in the rat. Involvement of skin in its catabolism

Iodinated recombinant human interleukin-6 (125I-rhIL-6) was intravenously injected into rats and its fate was studied during 24 h. Between 10-20 min after a single-dose injection, 125I-rhIL-6 accumulated in liver as previously reported [Castell et al. (1988) Eur. J. Biochem. 177, 357-361]. After 1 h, the radioactivity disappeared from the liver and accumulated in skin, reaching 35% of injected 125I-rhIL-6 5-8 h after injection. No comparable accumulation of radioactivity was found in skin when [125I]iodide or rat serum 125I-albumin was administered. Finally the radioactivity was detected as [125I]iodide in urine. Autoradiographic analysis of skin sections 5 h after 125I-rhIL-6 injection showed radioactivity in the interstitium. When the experiments were carried out with [35S]rhIL-6, essentially the same results were obtained: a decrease in radioactivity in the liver after 20 min, and a substantial increase in skin 7 h after injection. In vitro experiments showed that 125I-rhIL-6 is degraded by rat and human fibroblasts, whereas no degradation was observed with rat hepatoma cells (Fao) or human hepatocytes. These observations suggest the involvement of skin in the catabolism of IL-6.     

Prevalence of Mycobacterium tuberculosis infection among injection drug users in Toronto

BACKGROUND: Injection drug users are at increased risk of Mycobacterium tuberculosis infection and active tuberculosis (TB). The primary objective of this study was to determine the prevalence of M. tuberculosis infection among injection drug users in Toronto, as indicated by a positive tuberculin skin test result. An additional objective was to identify predictors of a positive skin test result in this population. METHODS: A cross-sectional study was carried out involving self-selected injection drug users in the city of Toronto. A total of 171 participants were recruited through a downtown Toronto needle-exchange program from June 1 to Oct. 31, 1996. RESULTS: Of 167 subjects tested, 155 (92.8%) returned for interpretation of their skin test result within the designated timeframe (48 to 72 hours). Using a 5-mm cut-off, the prevalence rate of positive tuberculin skin test results was 31.0% (95% confidence interval 23.8% to 38.9%). Birth outside of Canada and increasing age were both predictive of a positive result. INTERPRETATION: There is a high burden of M. tuberculosis infection in this population of injection drug users. The compliance observed with returning for interpretation of skin test results indicates that successful TB screening is possible among injection drug users.     

Critical ischemia times and survival patterns of experimental pig flaps

Previous work on critical ischemia time suggested (1) a greater susceptibility of myocutaneous flaps over skin flaps to the ischemia reperfusion injury and (2) that duration of ischemia may affect the survival area of a flap. Using a pig model, 55 animals were operated on and the critical ischemia times and survival patterns of the buttock skin (n = 85) and latissimus dorsi myocutaneous (n = 88) island flaps were determined after being submitted to 0, 2, 4, 6, 8, 10, 12, 14, and 16 hours of normothermic ischemia. The average critical ischemia times (CIT50) were determined to be 9 and 10 hours for the buttock skin and latissimus dorsi myocutaneous flaps, respectively. Percentage of total area surviving (%TAS) in those flaps which did survive was adversely affected by increases in the ischemic interval in both flap models. A statistically significant decrease in percentage of total area surviving was found after 6 and 8 hours of ischemia for the buttock skin and latissimus dorsi myocutaneous flaps, respectively.     

The role of allopurinol and deferoxamine in preventing pressure ulcers in pigs

Ischemia and reperfusion may be important in the pathogenesis of pressure ulcers. On the basis of this hypothesis, the effects of intermittent pressure and the anti-free radical agents allopurinol and deferoxamine were studied in a pig model in which a pressure of 150 mmHg was applied intermittently to the scapulae. Cutaneous blood flow, transcutaneous oxygen tension, skin and muscle damage, and muscle levels of adenosine triphosphate were quantified. A control group of pigs (n = 6) was untreated, the allopurinol group (n = 6) received oral allopurinol beginning 2 days before the experiment, and the deferoxamine group (n = 6) received an intramuscular injection of deferoxamine 2 hours before the experiment. Pressure (150 mmHg) was applied to the scapulae for 210 minutes, and it was relieved for 30 minutes. This 4-hour cycle was repeated continuously for 48 hours, and it resulted in pressure injuries in all animals. Allopurinol and deferoxamine improved cutaneous blood flow and tissue oxygenation, but only deferoxamine could significantly reduce cutaneous and skeletal muscle necrosis (p < 0.001). This study suggests a future role for anti-free radical agents in the reduction of pressure-induced injury.     

The critical relationship between free radicals and degrees of ischemia: evidence for tissue intolerance of marginal perfusion

Skin-flap ischemia has been associated with the presence of free radicals. In this study, two enzyme systems involved in free-radical metabolism were used to compare a distal skin flap to a skin graft. Forty-two rats were divided into several test groups. A 10 X 3 cm dorsal rat flap was used, and tissue biopsies for xanthine oxidase and malonyldialdehyde (MDA) were obtained 2.5, 5.5, and 8.5 cm from the base of the flap at the hours given. In group I (control), the flap was outlined but not elevated, and biopsies were obtained. In group II, the flap was elevated, and biopsies were obtained at 6 hours. In group III, the flap was elevated, the distal 4 X 3 cm was amputated and replaced as a full-thickness skin graft, and biopsies were obtained at 6 hours. In group IV, the flap was elevated, and biopsies were obtained at 12 hours. In group V, the flap was treated as in group III, and biopsies were obtained at 12 hours. In group VI, the flap was elevated, and biopsies were obtained at 24 hours. In group VII, the flap was treated as in group III, and biopsies were obtained at 24 hours. Results: Xanthine oxidase was significantly higher in all distal biopsies compared to proximal biopsies. Xanthine oxidase also increased with time. Malonyldialdehyde increased over time as well as with distance from the flap base. Distal flap biopsies at 24 hours had greatly increased levels of malonyldialdehyde compared to skin grafts (p less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)     

Experimental vaccinal prophylaxis of Pseudomonas aeruginosa burn sepsis

After the injection of P. aeruginosa live culture under the burned skin of mice sepsis develops within the first 24 hours, finally leading to the death of the animals. The microorganisms can be isolated from the blood, liver, kidneys and mesenterial lymph nodes till day 3 and from the spleen till day 5. After the intraperitoneal injection of P. aeruginosa live culture into mice, sepsis also develops within 24 hours, and the culture can be isolated from the blood and parenchymatous organs till day 3. The LD50 of the culture is equal to 5.1 X 10(6) microbial cells when introduced intraperitoneally and to 30 microbial cells in experimental burn sepsis. Experimental burn sepsis clearly demonstrates the effectiveness of Pseudomonas acellular protein vaccine: its index of effectiveness exceeds 3,000.     

Effects of sensitization of the guinea pig slow muscle in disorders of the neurotrophic control]

The immunohistochemical profile of intact and denervated soleus muscle of guinea pigs after sensibilization was studied. It is shown, that intact soleus muscle consists of slow fibers, which have low ATP-ase activity and don't react with monoclonal antibodies against fast myosin heavy chain. No changes of immunohistochemical profile were found after denervation or sensibilization. At the same time, the fibers, reacting with monoclonal antibodies against fast myosin heavy chain and having low ATP-ase activity, were found in denervated muscles after sensibilization. It is concluded, that the synthesis of fast myosin is induced after sensibilization of denervated muscles. Validity of myosin ATP-ase histochemistry for muscle fibers typing is discussed.     

Regulatory mechanisms of cutaneous delayed-type hypersensitivity. II. Suppression of cutaneous delayed-type hypersensitivity by macrophage disappearance reaction

Studies were performed on the behavior of cutaneous delayed-type hypersensitivity (DTH) in guinea pigs in which macrophage disappearance reaction (MDR) was induced. Guinea pigs were immunized with dinitrophenylated egg albumin (DNP-EA), followed by intraperitoneal (ip) injection of liquid paraffin in order to elicit peritoneal macrophages. Subsequently 20 micrograms of EA was injected into these animals and the animals were divided into two groups. One group of animals was sacrificed for estimation of MDR 6 hr after the subsequent ip injection. The other group received a skin test by EA at the time of the subsequent ip injection. The first group of animals sacrificed for estimation of MDR exhibited a marked reduction in the number of peritoneal macrophages. The second group of animals that received skin tests revealed suppressed skin reactions 24 hr after the subsequent ip injection. A similar experiment was performed using the guinea pigs doubly immunized with DNP-EA and dinitrophenylated bovine gamma-globulin (DNP-BGG). Induction of MDR was performed by ip injection of BGG and skin tests were done by both EA and BGG. As a result, suppression of not only BGG-induced skin reactions but also EA-induced skin reactions was observed in animals in which MDR had been induced by BGG. In addition, the guinea pigs in which MDR was induced showed hyporeactivity to phytohemagglutinin (PHA). Reactivity to skin reactive factor (SRF) was also suppressed in these animals. The culture supernatants of macrophages incubated with the MIF fraction in vitro showed the ability to suppress skin reactions of cutaneous DTH, PHA and SRF.     

Effects of indomethacin and prostaglandins on ovulation of goldfish.

A technique is described whereby elevated temperature and HCG injection yield a high percentage of ovulation in gravid goldfish. Indomethacin (10 mug/g; i.p. injection) completely inhibits ovulation if given within 6 hours following HCG (4 IU/g); the unovulated oocytes develop rapidly into corpora atretica. PGE1, PGE2, and PGF2alpha (5 mug/g; i.p. injection) induce ovulation in fish treated with indomethacin and HCG; PGE2 was most effective when given 11 hours after HCG. The results suggest that the ovulatory action of prostaglandins following HCG stimulation is at the level of the ovary and that it is restricted to a period between 7 and 12 hours after the gonadotropin injection.     

Mouse splenocyte sensitization to normal tissue antigens and natural killer activity. III. The effect of partial hepatectomy

Partial hepatectomy leads to both increasing of natural cell-mediated activity and sensibilization level (SL) of splenocytes of hepatectomized mice towards antigens of the syngeneic liver. The wave-like variability of SL was shown with sharp increase at 3, 6 and 9 days after operation. Natural killer activity was elevated on the 2nd and 10th days with a significant decrease on the 3-4th days after operation. It is assumed that the variability in the functional activity of splenocytes under study may characterized splenocytes of different populations.     

Induction of C-reactive protein, serum amyloid P component, and kininogens in the submandibular and lacrimal glands of rats with experimentally induced inflammation.

The mRNAs for acute-phase proteins and kininogens were found to be increased in the submandibular gland (SMG) and extraorbital and intraorbital lacrimal gland (ELG and ILG) in response to experimentally induced inflammation in rats; i.e., 24 hours after subcutaneous injection of turpentine oil, mRNAs for C-reactive protein (CRP), serum amyloid P component (SAP), and H- and T-kininogens were induced in the SMG, ELG, and ILG of rats, whereas these mRNAs were not detected in the same tissues of normal control rats. The induction of mRNAs for these inflammatory proteins by turpentine oil was preceded by a transient increase in the level of mRNA for tumor necrosis factor-alpha (TNF-alpha) at 6 hours after subcutaneous injection of the oil. This was confirmed by injection of another inflammation inducer, lipopolysaccharide (LPS), which induced the TNF-alpha mRNA in the same way at 6 hours as turpentine oil did. The up-regulation of acute-phase proteins including kininogens in the SMG, ELG, and ILG suggest the existence of a strict defense system in the exocrine glands.     

Effect of tuftsin on enzyme activity in the energy metabolism of lymphocytes

The cytochemical technique was used to measure the activity of succinate dehydrogenase (SDH), lactate dehydrogenase (LDH) and glucose-6-phosphate dehydrogenase (G-6-PDH) of peripheral blood lymphocytes of mice and rats given intraperitoneal injections of an endogenous immunostimulant tuftcin (Tre-Lys-Pro-Arg) in a dose of 0.3 mg/kg. A significant decrease of SDH activity was observed both in mice and rats 4 and 6 hours following injection, respectively. In mice, that activity returned to normal in 12, while in rats in 24 hours. An opposite action was produced by tuftcin on G-6-PDH, causing the maximum elevation of the enzyme activity in rat lymphocytes 6 hours after peptide administration. The decrease to the initial level was observed in 24 hours. Tuftcin did not affect the activity of LDH. The data obtained indicate that the immunological effect of tuftcin is coupled with the changes in the activity of Krebs cycle enzymes (SDH) and pentose phosphate cycle enzymes (G-6-PDH).