系统性光变态反应检测动物模型的建立

目的建立稳定、完善的系统性光变态反应检测的动物模型。方法以对氨基苯磺酰胺（SN）作为光敏剂,经腹腔注射与灌胃两种途径对小鼠和豚鼠进行给药,通过UVA加UVB光照诱导以及只用UVA光照激发后,于4h、24h、48h和72h分别测量小鼠耳厚度及观察豚鼠皮肤红斑反应。结果24h、48h和72h时,小鼠出现耳肿胀反应,豚鼠出现皮肤红斑反应。结论口服、腹腔注射SN,均可产生光变态反应,成功建立系统性光变态反应检测的动物模型。     

Scaled-up production and purification of Clostridium perfringens type A enterotoxin

Methods for small-scale production of Clostridium perfringens type A enterotoxin were unsuitable for large-scale culture of this organism. Rapid, efficient harvesting of 40 1 batch culture of Cl. perfringens was achieved by tangential flow micro-filtration with the Millipore Pellicon cassette system. Enterotoxin-containing extracts were prepared by passing concentrated suspensions of the harvested cells through a French pressure cell. The overall yield of purified enterotoxin was 38·8%. The toxin gave a single band on native polyacrylamide gels but formed high molecular weight aggregates in the presence of sodium dodecyl sulphate. These aggregates frequently occurred during storage of non-sterile enterotoxin preparations but could be separated from the monomer toxin by gel filtration on Sephadex G-100. Purified monomer enterotoxin had biological activities of 119·3 μ g/kg mouse lethal dose when injected intraperitoneally and 3333 capillary permeability increasing units/mg protein in guinea pig skin. Thirty μg of the enterotoxin caused fluid accumulation in ligated rabbit ileal loops. Aggregated enterotoxin had no demonstrable biological or immunological activity.     

Scaled-up production and purification of Clostridium perfringens type A enterotoxin

Methods for small-scale production of Clostridium perfringens type A enterotoxin were unsuitable for large-scale culture of this organism. Rapid, efficient harvesting of 40 1 batch culture of Cl. perfringens was achieved by tangential flow micro-filtration with the Millipore Pellicon cassette system. Enterotoxin-containing extracts were prepared by passing concentrated suspensions of the harvested cells through a French pressure cell. The overall yield of purified enterotoxin was 38.8%. The toxin gave a single band on native polyacrylamide gels but formed high molecular weight aggregates in the presence of sodium dodecyl sulphate. These aggregates frequently occurred during storage of non-sterile enterotoxin preparations but could be separated from the monomer toxin by gel filtration on Sephadex G-100. Purified monomer enterotoxin had biological activities of 119.3 micrograms/kg mouse lethal dose when injected intraperitoneally and 3333 capillary permeability increasing units/mg protein in guinea pig skin. Thirty micrograms of the enterotoxin caused fluid accumulation in ligated rabbit ileal loops. Aggregated enterotoxin had no demonstrable biological or immunological activity.     

Heat-labile toxin-producing isolates of Pasteurella multocida from rabbits.

Five of one hundred forty seven isolates of Pasteurella multocida from rabbits were found to produce heat-labile toxin. Each isolate was assayed for the ability of potassium thiocyanate (KSCN) extracts to cause dermonecrosis in guinea pig skin, ability of bacteria or filtrates to cause cytotoxicity in cell cultures, and reactivity with monoclonal antibodies to heat-labile P. multocida toxin. Five capsular type D isolates produced dermonecrosis and reacted with monoclonal antibodies to toxin. Filtrates of all five of these isolates were cytotoxic for cell cultures. Potassium thiocyanate extracts of all five isolates caused pleuritis and pneumonia in rabbits after intranasal inoculation. Turbinate atrophy was seen in 5 of 19 rabbits inoculated intranasally with toxic extracts. Heat-labile toxin was not produced by 109 capsular type A isolates or 19 nontypable isolates.     

Factors influencing the toxicity of salivary gland extracts of Ixodes holocyclus Neumann

A bioassay using mice was developed to compare the toxin content of extracts of salivary glands of I. holocyclus at various stages of feeding. The quantity of toxin increased rapidly from the third day of feeding. Toxin production continued and increased in ticks removed after 3–5 days on mice and held at 30°C at 92% RH for 24 h, whereas no toxin was detected in the salivary glands of ticks fed for 3 days and treated similarly. It is suggested that major physiological changes occur in the salivary glands of I. holocyclus on the third day, which once stimulated continue independently of feeding. Toxin production in ticks was not suppressed by passively immunizing mice with anti-tick toxin but was in ticks fed upon hosts with a previous experience of tick feeding.Thus, to obtain salivary glands containing high concentrations of toxin for chemical analysis, it is necessary for salivary glands to develop 5 days from the initial attachment of the tick to a host with no previous experience of tick feeding. This can be achieved by passively immunizing mice against toxin, thus enabling the tick to feed 5 days without killing the mouse or by keeping the tick for 24 h at 30°C at 92% RH following the death of the mouse on the fourth day.     

The detection of the messenger ribonucleic acid for the α-lactalbumin of guinea-pig milk (Short Communication)

RNA was extracted from the polyribosomes isolated from the mammary glands of a lactating guinea pig and injected into Xenopus oocytes. On incubation the oocytes effected the biosynthesis of alpha-lactalbumin.     

Evaluation of a guinea pig model to assess interference in the immunogenicity of different components of a combination vaccine comprising diphtheria, tetanus and acellular pertussis (DTaP) vaccine and haemophilus influenzae type b capsular polysaccharide conjugate vaccine.

A guinea pig model to assess the immunogenicity of a combination vaccine containing diphtheria, tetanus and acellular pertussis (DTaP) vaccine and Haemophilus influenzae type b (Hib) capsular polysaccharide conjugated to tetanus toxoid (HibT) was evaluated comparatively with the mouse immunogenicity test to study the effect of combining these antigens on the immunogenicity of various components. The immunogenicity test in mice was performed by subcutaneous injection of groups of 10 animals twice at an interval of four weeks with 1/10 of a single human dose of various formulations of combination vaccines, DTaP or HibT vaccine. The animals were bled at 4 and 6 weeks and IgG or total antibodies to various components were determined by ELISA or RIA. The guinea pig immunogenicity model included groups of animals injected subcutaneously twice at an interval of six weeks with 1.5 times the single human dose of various formulations. The animals were bled at 4, 6 and 8 weeks and serum samples were tested for antibodies to various components by ELISA, RIA and/or neutralization tests. Additionally, potency of tetanus and diphtheria components was assessed as per the US Food and Drug Administration's regulations. Aluminium phosphate (AIPO(4)) adsorbed HibT vaccine or HibT as a combination with AIPO(4)adsorbed DTaP vaccine showed significant increases in IgG antibodies to tetanus toxin in mice as well increased tetanus antitoxin levels in guinea pigs as compared to soluble HibT vaccine. In general, combining DTaP and HibT vaccines did not affect the antibody levels to tetanus and diphtheria toxoids whereas DTaP-HibT combination vaccine elicited significantly lower IgG antibodies to pertussis toxin and filamentous haemagglutinin than DTaP vaccine alone, particularly after first injection. Mice showed similar Hib antibody responses for the combination and HibT alone whereas guinea pigs consistently showed lower anamnestic responses to Hib for combination formulations than for HibT alone. Reducing the amount of HibT and/or tetanus toxoid in the combination formulations reduced this suppression of Hib antibody response in guinea pigs. Suppression of Hib antibody response in combination vaccines has also been reported from recent clinical trials. Based on the results from this study, it appears that the guinea pig model may be able to predict the human response to various components of combination vaccines.     

Pharmacological studies on the venomous spotted butterfish (Scatophagus argus Linn) sting extract on experimental animals

A sting of the fish S. argus, a venomous edible spotted butterfish, produces tremendous local pain, severe swelling, rise of body temperature, throbbing sensation etc. To establish the pharmacological activities of S. argus sting extract, the present investigation, was carried out on experimental animals. The LD50 of extract was found to be 9.3 mg/kg (iv) in male albino mice. The extract showed loss of sensation, urination and salivation in mice. It potentiated pentobarbitone induced sleeping time in male albino mice and produced hypothermia. Extract produced a fall of cat and guinea pig blood pressure, which was completely abolished by mepyramine. It produced a transient reduction of respiratory rate in rat, but decreased respiratory amplitude in cat, which was abolished after vagotomy. On isolated toad heart, the extract increased both the amplitude and rate of contraction. On isolated guinea pig heart, the sting extract decreased both the rate and amplitude of contraction leading to cardiac arrest, but it had no effect on isolated guinea pig auricle. The extract produced a reversible blockade of electrically induced twitch response of isolated chick biventer cervices preparation, but it had no effect on the isolated rat phrenic nerve diaphragm preparation. It produced a slow contractile response on isolated guinea pig ileum, rat uterus and rat fundal strip preparations but produced slow relaxation on isolated rat duodenum preparation. The contractile response on isolated guinea pig ileum and rat fundal strip was antagonised by SC19220. It did not produce any significant cutaneous haemorrhage in mice and did not produce any haemolysis on saline washed erythrocytes. The sting extract significantly increased capillary permeability of guinea pig dorsal flank and produced oedema in mice hind paw.     

Detection of Clostridial Hemolysin Formed In Vivo

Experimental clostridial myonecrosis closely resembling gas gangrene was produced in goats by the intramuscular injection of washed clostridia suspended in adrenalin. The preparation provides a model for the study of clostridial myonecrosis that is relatively isolated from preceding trauma. A new test for the demonstration of clostridial exotoxins, the hemolysin indicator plate test, was described. This test gave reliable results within 6 hr, and only minute quantities of test material were required. Application of this test to would exudates of goats with experimental clostridial myonecrosis demonstrated the presence of the gamma toxin of Clostridium novyi, alpha toxin of C. septicum, and the alpha toxin of C. perfringens in exudates from animals injected with washed Clostridium bacilli suspended in adrenalin. Exudates from goats injected with C. perfringens were lethal for mice and produced necrosis of guinea pig skin typical of C. perfringens. The convincing demonstration of clostridial exotoxins formed in vivo in this experimental model could be the basis for definition of clostridial exotoxin actions which have evaded study in the past.     

Effect of methylglyoxal bis (guanylhydrazone) (MGBG) in vivo on the decarboxylation of s-adenosylmethionine and synthesis of spermidine in the rat and guinea pig.

The rates of decarboxylation of $\text{S}$-adenosylmethionine and synthesis of spermidine have been measured in extracts of liver, kidney and brain of the rat and guinea pig after intraperitoneal injection of MGBG, both before and after dialysis. The rate of decarboxylation of $\text{S}$-adenosylmethionine paralleled that of spermidine synthesis in all of the tissues investigated, even when spermidine synthesis was measured using preformed decarboxylated $\text{S}$-adenosylmethionine as substrate instead of $\text{S}$-adenosylmethionine itself. MGBG inhibited CO₂ production and spermidine synthesis to a similar extent in extracts of liver and kidney of both the rat and the guinea pig. After dialysis, a similar increase in both CO₂ production and spermidine synthesis was noted in these extracts. No effects on CO₂ production or spermidine synthesis were noted on extracts of brain of the rat or guinea pig, either before or after dialysis. When MGBG was injected intracisternally, CO₂ production and spermidine synthesis by extracts of brain were inhibited to the same extent, and after dialysis a similar increase in CO₂ production and spermidine synthesis was observed. These results indicate that the effects of MGBG are essentially the same in brain as they are in liver and kidney, and the MGBG injected intraperitoneally does not pass into the brain.     

Molecular cloning and physical mapping of the hyd gene of Escherichia coli K-12

Abstract Passive transfer between rates of protection against cholera toxin (CT) was studied. Extracts of various organs, obtained from CT-immunized rats, were injected intravenously into non-immunized recipient rats. The ability of the extracts to inhibit CT-induced secretion in ligated jejunal loop were tested. A significant inhibition of the response to CT was achieved by extracts from hypophysis, brain and jejunal mucosa. Extracts from pancreas, spleen or adrenal glands were without effect, as were all extracts obtained from control rats. The antisecretory effects of the hypophysis extracts became intensified with increasing numbers of immunizations, and the antisecretory effect was most pronounced when the extract was injected immediately before the CT challenge. The active component of the hypophysis extract was heat-labile and negatively charged, suggesting an acidic protein as the mediator of the protective effect against CT.     

Alpha-adrenergic receptor blocking effect of Cleistanthus collinus (Roxb.) Benth. and Hook f. leaf extract on guinea pig isolated smooth muscle preparations

Aqueous extract of C. collinus leaves inhibited norepinephrine induced contraction in guinea pig vas deferens and aortic strip in a dose-dependent manner. Inhibition of acetylcholine induced contraction in ileum was dose independent. C. collinus extract per se had no effect on isolated guinea pig vas deferens and aortic strip, but inhibited norepinephrine induced contraction in a dose-dependent manner probably by its antagonist action on alpha-adrenergic receptor. It had inconsistent effect on guinea pig ileum in vitro preparation.     

Fluoride-mediated activation of guinea pig neutrophils

In guinea pig periotoneal neutrophils NaF at a concentration of above 5 mM elicited a dose-dependent, delayed and sustained activation of NADPH oxidase. Unlike in human neutrophils, in guinea pig cells, this response was independent of extracellular calcium. Fura2 fluorescence measurements indicated also a fluoride-mediated moderate elevation in the level of cytosolic calcium concentration. Pretreatment of neutrophils with pertussis toxin, blocked fluoride-promoted activation of NADPH oxidase, indicating that NaF stimulation was mediated by a G protein which is a pertussis toxin substrate. NaF-elicited calcium elevation was insensitive to the toxin. Upon transfer of NaF-stimulated cells to a fluoride-free medium, superoxide release declined and calcium levels diminished. The response of the deactivated, fluoride-prestimulated guinea pig neutrophils to a secondary stimulation with phorbol myristate acetate (PMA) or fMet-Leu-Phe, was either unaffected by the previous challenge with NaF (PMA) or augmented by it (the chemotactic peptide). In parallel to the activation of NADPH oxidase, NaF also induced translocation of protein kinase C to cell membranes. This effect was also abolished by a pretreatment with pertussis toxin.     

Biological activities of a chemically synthesized form of leukotriene E4

A chemically synthesized form of leukotriene E₄ (LTE₄) has been studied for its ability to induce contractions in isolated guinea pig ilea, to induce vascular permeability changes in rat skin when injected intradermally, and to induce bronchoconstriction in guinea pigs after intravenous injection. The synthetic compound induced a contraction in the guinea pig ileum which was slower in developing than that induced by histamine but faster in developing than that induced by a crude preparation of SRS-A isolated from guinea pig lung. The compound was 70-fold more active than histamine on the guinea pig ileum (EC₅₀ of 5 × 10^?9 and 3.5 × 10^?7 M, respectively). FPL 55712, a known SRS-A antagonist, exhibited the same potency in blocking the contractions elicited by the synthetic material as it did in blocking contractions produced by guinea pig SRS-A generated biologically (IC₅₀ of 3.5 × 10^?8 M). The synthetic LTE₄ induced a dose dependent increase in vascular permeability in the rat skin which was antagonized by the intravenous injection of FPL 55712 (ID₅₀ of 1.2 mg/kg). The synthetic material was also a potent bronchoconstrictor in the guinea pig when injected intravenously. The bronchoconstriction, too, was antagonized by FPL 55712 when injected intravenously (ID₅₀ of 0.2 mg/kg). In both the rat and guinea pig, FPL 55712 exhibited a short duration of action $\text{in vivo}$. The $\text{in vivo}$ model systems discussed in this study, utilizing the synthetic form of LTE₄ should be useful in the future evaluation of other SRS-A antagonists.     

Winter accumulation of paralytic shellfish toxins in digestive glands of mussels from Arcachon and Toulon (France) without detectable toxic plankton species revealed by interference in the mouse bioassay for lipophilic toxins

Since January 1993, neurological symptoms and rapid deaths (5 to 10 min) were typically observed in the mouse bioassay of acetone extracts of digestive glands from Arcachon and Toulon (France) during the winter season. It was assumed initially that a new lipophilic toxin was present because tests using the AOAC mouse bioassay for paralytic shellfish toxins on acid extracts of whole shellfish meat were negative, no known lipophilic toxins were detected and no toxic phytoplankton species were observed in the area during the poisoning events. In this study, however, preparative isolation of the toxic factor from toxic mussel digestive glands has revealed the presence of paralytic shellfish toxins, the principal ones being gonyautoxins-2 and -3 at Arcachon and gonyautoxins-1, -4, -2 and -3 at Toulon. The toxin concentrations recorded were below levels harmful to consumers and therefore represent a false positive in the mouse bioassay for lipophilic toxins based upon acetone extraction. The origin of the toxins remains to be determined.     

Fluoride-mediated activation of guinea pig neutrophils

In guinea pig peritoneal neutrophils NaF at a concentration of above 5 mM elicited a dose-dependent, delayed and sustained activation of NADPH oxidase. Unlike in human neutrophils, in guinea pig cells, this response was independent of extracellular calcium. Fura2 fluorescence measurements indicated also a fluoride-mediated moderate elevation in the level of cytosolic calcium concentration. Pretreatment of neutrophils with pertussis toxin, blocked fluoride-promoted activation of NADPH oxidase, indicating that NaF stimulation was mediated by a G protein which is a pertussis toxin substrate. NaF-elicited calcium elevation was insensitive to the toxin. Upon transfer of NaF-stimulated cells to a fluoride-free medium, superoxide release declined and calcium levels diminished. The response of the deactivated, fluoride-prestimulated guinea pig neutrophils to a secondary stimulation with phorbol myristate acetate (PMA) or fMet-Leu-Phe, was either unaffected by the previous challenge with NaF (PMA) or augmented by it (the chemotactic peptide). In parallel to the activation of NADPH oxidase, NaF also induced translocation of protein kinase C to cell membranes. This effect was also abolished by a pretreatment with pertussis toxin.     

Lethal and Dermonecrotic Activities of Clostridium perfringens Iota Toxin: Biological Activities Induced by Cooperation of Two Nonlinked Components

The effect of separate injections of two components of Clostridium perfringens iota toxin, designated I_a and I_b components, on the biological activities of the toxin was investigated. The intravenous injection of one component within 120 min after the injection of the other component killed mice. The activity of iota toxin was abolished by anti-I_a or anti-I_b antiserum. On the other hand, when I_b component was intravenously administered to mice given anti-I_a antiserum within 120 min after the intravenous injection of I_a component, the lethal activity was completely neutralized, but when I_a component was injected into mice that were given anti-I_b antiserum over 5 min after the injection of I_b component, the activity was not neutralized. The separate injections of I_a and I_b components in skin of guinea pigs indicated dermonecrosis at the injection site of I_b components, but not at the site of I_a components. Furthermore, when one component was intradermally injected in guinea pigs and then the other intraperitoneally, the dermonecrotic activity of the toxin was observed at the intradermal injection site of I_b component, but not at that of I_a component. From the data, it appears that the lethal and dermonecrotic activities of iota toxin are initiated by the binding of I_b component to specific sites on tissues.     

Notes on Technic: Differential staining of Fungus and host cells Using a modifciation of Pianeze IIIB

Intercellular secretory capillaries in parotid glands, eccrine sweat glands and intracellular secretory capillaries in parietal cells of gastric glands were demonstrated histo-chemically by the use of the Wachstein-Meisel adenosinetriphosphatase (ATPase) technique in the rabbit, rat and guinea pig. However, with the Wachstein-Meisel 5-nucleotidase technique, secretory capillaries were not stained. For parotid glands, optimal incubation in ATPase substrate mixture was: in rabbit, 15 min; in rat, 2.5 hr; and in guinea pig, 2 hr. For eccrine sweat glands, optimal incubation was 15 min in rabbit, 30 min in rat and 15 min in guinea pig. For parietal cells of gastric glands, optimal incubation was 3 hr for all three species. Secretory capillaries were best demonstrated in the parotid by using rabbit tissue; in eccrine sweat glands, with rat tissue, and in parietal cells, guinea pig tissue. Since ATPase activity in cell membranes of secretory cells may play a part in the mechanism of transport of secretory products from their place of formation in the acini to the excretory ducts, the Wachstein-Meisel ATPase technique can therefore be used successfully for staining secretory capillaries in many of the exocrine glands of laboratory mammals.     

Staining Secretory Capillaries of Exocrine Glands with Techniques for Specific Phosphatases

Intercellular secretory capillaries in parotid glands, eccrine sweat glands and intracellular secretory capillaries in parietal cells of gastric glands were demonstrated histo-chemically by the use of the Wachstein-Meisel adenosinetriphosphatase (ATPase) technique in the rabbit, rat and guinea pig. However, with the Wachstein-Meisel 5-nucleotidase technique, secretory capillaries were not stained. For parotid glands, optimal incubation in ATPase substrate mixture was: in rabbit, 15 min; in rat, 2.5 hr; and in guinea pig, 2 hr. For eccrine sweat glands, optimal incubation was 15 min in rabbit, 30 min in rat and 15 min in guinea pig. For parietal cells of gastric glands, optimal incubation was 3 hr for all three species. Secretory capillaries were best demonstrated in the parotid by using rabbit tissue; in eccrine sweat glands, with rat tissue, and in parietal cells, guinea pig tissue. Since ATPase activity in cell membranes of secretory cells may play a part in the mechanism of transport of secretory products from their place of formation in the acini to the excretory ducts, the Wachstein-Meisel ATPase technique can therefore be used successfully for staining secretory capillaries in many of the exocrine glands of laboratory mammals.     

Complement-mediated alteration of antibody specificity in vivo.

The simultaneous injection of heterologous anti-EL4 lymphoma serum and complement results in the rapid disappearance of such antibody from the periphery of non-tumor bearing mice. However, this phenomenon is only observed when a complement source capable of mediating the lysis of EL4 cells sensitized with such heterologous antibody is used. This complement mediated enhancement of anti-tumor antibody absorption was observed in vivo for three strains of mice. Omission of complement or the use of genetically deficient complement sources resulted in no effect on circulating antibody titer when compared to the titer of heterologous anti-tumor antibody observed in the periphery when injected alone. Exogenous complement did not enhance the clearance of heterologous anti-tetanus toxin serum, thereby suggesting that the increased absorption of anti-EL4 in vivo is not related simply to the enhanced clearance of foreign gamma-globulin. Confirmatory evidence of the role of complement in altering anti-tumor antibody specificity in vivo was obtained in a guinea pig tumor model as well. The data suggest that anti-tumor serum shown to be relatively specific for the tumor cell gains additional specificity in the presence of functional complement and consequently manifests avidity for cross-reactive determinants previously thought to be unrelated.