Intraperitoneal Injection Of Cyclophosphamide Research Articles

The concepts of tail moment and tail inertia in the single cell gel electrophoresis assay.

Single cell gel electrophoresis under alkaline conditions is a technique used to detect primary DNA damage in individual mammalian cells. Cells embedded in agarose on microscope slides are subjected to lysis, unwinding of DNA and electrophoresis at high pH. After staining with a fluorescent dye, cells with DNA damage display increased migration of genetic material from the cell nucleus. The damage is quantified by measuring the displacement between the genetic material of the nucleus ('comet head') and the resulting 'tail'. The torsional moment of the tail ('tail moment') has been suggested to be an appropriate index of induced DNA damage in considering both the migration of the genetic material as well as the relative amount of DNA in the tail. In the present paper it will be shown that the moment of inertia ('tail inertia'), a not previously described tail parameter, provides a more precise description of the distribution of individual DNA fragments within the tails. The tail inertia was also found to be the most sensitive indicator of the DNA damage induced in peripheral lymphocytes from mice given a single intraperitoneal injection of cyclophosphamide (150 mg/kg b.w.). It is concluded that the tail inertia is an important complement to other tail parameters when looking for damage of DNA with the single cell gel electrophoresis assay.

In vivo administration of taurine and niacin modulate cyclophosphamide-induced lung injury

The antiinflammatory, antioxidant activity of taurine and niacin against cyclophosphamide-induced early lung injury in rats was investigated. A single intraperitoneal injection of cyclophosphamide markedly altered the levels of several biomarkers in bronchoalveolar lavage fluid: total protein, albumin, angiotensin converting enzyme, lactate dehydrogenase, lactate, N-acetyl- β- d-glucosaminidase, alkaline phosphatase, acid phosphatase and lipid peroxidation product were significantly elevated. In contrast, decreased levels of total reduced glutathione (GSH) and ascorbic acid were observed. Cyclophosphamide significantly increased malondialdehyde levels in serum and lung. Significant increases in lung content of lipid hydroperoxides were seen that paralleled the decreased levels of total reduced glutathione and total sulfhydryl groups. Pretreatment of rats with daily intraperitoneal injection of taurine plus niacin 7 days prior to and 2 days after cyclophosphamide insult significantly inhibited the development of lung injury, prevented the alterations in lavage fluid biomarkers associated with inflammatory reactions, with less lipid peroxidation and restoration of antioxidants. In conclusion, our results suggest that taurine and niacin in combination is efficient in blunting cyclophosphamide-induced pulmonary damage.

The regulation of intestinal hypersensitivity reactions to ovalbumin by omega-3 fatty acid enriched diet: studies of IEL and LPL in mucosal damage.

In order to clarify the mechanisms of food-sensitive enteropathy, a food hypersensitive model was generated by feeding ovalbumin to female BALB/c mice after intraperitoneal injection of cyclophosphamide and morphological and immunological changes in the gut mucosa were investigated. Villus atrophy, crypt hyperplasia and increased numbers of intra-epithelial lymphocytes (IEL) were confirmed in this model, as seen in food-sensitive enteropathy in humans. Subpopulations of IEL and lamina propria lymphocytes were enumerated by immunohistochemical observation. CD8-positive cells were increased both in epithelium and lamina propria, whereas CD4-positive cells were decreased in lamina propria. We document here that orally administered food antigen actually induces food-sensitive enteropathy and mucosal damage is generated by lymphocytes that infiltrate the intestinal mucosa. We also investigated the effect of feeding an omega-3 fatty acid-enriched diet in this model and found that it was efficient in attenuating mucosal damage.

Cyclophosphamide induced early biochemical changes in lung lavage fluid and alterations in lavage cell function.

The present investigation evaluated the changes in bronchoalveolar lavage fluid (BALF) biochemical constituents and indices of bronchoalveolar lavage cell functions to detect early lung injury in rats following intraperitoneal administration of cyclophosphamide (CP). Rats were exposed to a single intraperitoneal injection of CP (200 or 300 mg/kg body weight). Experimental and control rats were sacrificed at various time intervals (2, 3, 5, 7, 11, 21, and 42 days after cessation of exposure), and lung lavage was performed to examine several markers of lung injury. Biochemical analyses revealed dose-related increases in BALF angiotensin converting enzyme activity, total protein, lactate, lactate dehydrogenase, and N-acetyl-beta-D-glucosaminidase (NAG) levels on days 2, 3, 5, 7, and dose-related increases in albumin, alkaline phosphatase, acid phosphatase, and lipid peroxidation on days 2, 3, 5, 7, and 11 after CP treatment. In contrast, reduced levels of ascorbic acid and glutathione (GSH) content were observed in lung lavage fluid. We also examined bronchoalveolar lavage cells for acid hydrolases (acid phosphatase, beta-glucuronidase, NAG) and GSH content. Activity of acid hydrolases was slightly elevated on day 2 and peaked on days 3, 5, and 7. However, lavage cell GSH content was decreased. Thus, measurements of pulmonary changes by analyzing lavage fluid and lavage cell functions seems to be a useful marker for assessing the early onset and development of CP-induced lung injury.

Gastrointestinal invasion by herpes simplex virus type 1 inoculated cutaneously into the immunosuppressed mice.

The pathogenesis of infection in mice with herpes simplex virus type 1 (HSV-1) strain 7401H was studied. Mice immunosuppressed by intraperitoneal injection of cyclophosphamide were inoculated cutaneously into the flank with the virus and developed severe zosteriform skin lesions. All of them died within 2 weeks after the infection, while most of the normal mice survived the viral infection with healing of the lesions. In the gastrointestinal tract of the immunosuppressed mice, macroscopic abnormalities were frequently observed, and infectious viruses were detected on days 7 to 9. The viruses were also detectable in the dorsal root ganglia and the spinal cord of thoracolumbar area on days 5 to 7, and in the celiac plexus on day 7. However, no viruses were detected in the blood. Immunohistological examination of the gastrointestinal tract revealed that the viral antigens were localized in Auerbach's myenteric plexus. These results suggest that HSV-1 inoculated into the flank skin invaded the gastrointestinal tract via the nervous system, which gastrointestinal involvement might possibly have caused the death of the mice.

Role of leukocyte depletion in noncholinergic bronchoconstriction of guinea pigs

To test the role of leukocytes in the activation of afferent C-fibers in the lung, 33 guinea pigs, 18 control and 15 experimental or leukocyte depleted, were used. The leukocyte depletion was accomplished with an intraperitoneal injection of cyclophosphamide (100 mg/kg) 96 h prior to the study. On the day of the study, bronchial constriction was produced either by exsanguination ( n = 17) or by capsaicin injection (16 μg/kg, i.v.) ( n = 16) in anesthetized-paralyzed animals. Venous blood samples were collected for leukocyte counts. At 1–30 min following the above treatment, the maximal expiratory flow maneuver was performed and a decrease in the maximal expiratory flow at 50% baseline total lung capacity (żmax 50) was used as an index of bronchoconstriction. The leukocyte count decreased significantly following the pretreatment with cyclophosphamide [an average of 6217 ± 612 (control) vs. 2242 ± 334/mm 3 (experimental)]. Exsanguination caused a gradual decrease in V̇max 50 with time, indicating a temporal increase in bronchoconstriction. Capsaicin injection, on the other hand, caused an immediate (1 min) marked bronchoconstriction, which attenuated gradually with time. At a specific time point, leukocyte depletion did not produce any significant change in V̇max 50 compared to the control group for both types of bronchoconstrictions. Based on these results, we conclude that leukocytes may play an insignificant role in the bronchoconstriction caused by the activation of afferent C-fibers in guinea pig lungs under our experimental conditions.

Contact sensitivity to captafol in BALB/c mice

The fungicide captafol has been reported as causing irritant and allergic contact dermatitis in humans and in guinea pigs. This study investigated the ability of purified captafol to cause contact sensitization in BALB/c mice. Female mice were pretreated with an intraperitoneal injection of cyclophosphamide or saline. Applications of captafol (18.7 or 37.4 mg/ml), dinitrofluorobenzene (DNFB; 5 mg/ml) or solvent (4:1 acetone:ethanol mixture) were administered to the shaved abdomen on 2 consecutive days, or on days 1, 2, 8, 15, 22 and 29. Following challenge with captafol (37.4 mg/ml) or DNFB (2 mg/ml) on the right ear 6 days after the last induction, ear thickness ratios (right ear/left ear) were significantly larger after challenge in captafol-induced and DNFB-induced mice compared to control mice. A slightly larger response was observed with the smaller induction dose level of captafol and with multiple inductions over the course of a month. The overall maximum response to captafol was not increased by pretreatment with cyclophosphamide. Histologically, ears from captafol-induced and DNFB-induced mice showed edema and cellular infiltration. This study demonstrated the ability of captafol to produce contact hypersensitivity in the BALB/c mouse.

Evaluation of antibody-bearing liposomal amphotericin B in the treatment of systemic candidiasis in a neutropenic murine model

The efficacy of liposomal amphotericin B bearing anticandidal antibodies (LAMB-Ab) was investigated in the treatment of systemic candidiasis in a murine model made neutropenic by an intraperitoneal injection of cyclophosphamide. Treatment with a single dose (0.6 mg amphotericin B kg-1 body weight) of LAMB-Ab resulted in an improved survival of neutropenic mice infected with Candida albicans compared to neutropenic mice treated with identical doses of liposomal amphotericin B or free amphotericin B.

Reproducibility of Seasonal Effects in Antibody Formation and Host Resistance in the Outbred Mouse

The purpose of this study was to assess the reproducibility of any seasonal effects in the outbred CD1 mouse of antibody production to sheep red blood cells (SRBCs) and host resistance to the bacterium Listeria monocytogenes. A marked seasonal effect on antibody production was seen when 5- to 6-week-old female CD1 mice were studied on a weekly basis for a period of 2 years. Maintained on a 12:12 h light:dark schedule, animals were held for 12 days prior to experiment to insure physical condition and acclimatization to the lighting regimen. Beginning at 4 h after lights on (HALO) for day 1 and 2 HALO thereafter, groups of mice were (a) not treated, (b) administered a vehicle (corn oil, 1% methylcellulose, or distilled water) by oral gavage for 5 days, or (c) not treated, but given an intraperitoneal injection of cyclophosphamide 24 h prior to assay. On the fifth day, mice were injected with SRBCs intravenously. Four days later, antibody formation against SRBCs was measured using spleen cells. Circannual and seasonal rhythms were displayed by each group of animals, with greatest antibody production, indicated by the number of plaque-forming cells (PFCs)/million viable cells, in the Spring (range of double amplitude = 36-108%). The timing of these rhythms was reproducible from one year to the next. In contrast, the magnitude of the response in year 1 was significantly different from year 2 for animals given vehicle or not treated. Cyclophosphamide-treated mice had consistently low numbers of plaque-forming cells. Host resistance was studied in separate mice treated with vehicles at the same time as the antibody assay. On the third day of dosing, mice were injected intravenously with Listeria monocytogenes (LM) and monitored for death for 10 days. When analyzed by Kruskal-Wallis life table analysis, there was no overall effect of vehicle on survival for 1987 but there was an effect for 1988 and when data from both years were combined. Distilled water-treated mice had lower survival rates than the other two vehicles. Mice treated with distilled water displayed a circannual rhythm for survival for 1988 and for both years combined, in contrast to the other two vehicles. When each vehicle was analyzed separately for seasonal effect, a significant effect of season occurred for corn oil- and distilled water-treated animals. The greatest survival rate and longest survival time generally occurred in the months between July and December.

Effects of Cyclophosphamide on the Pathogenesis of Cytomegalovirus-Induced Labyrinthitis

Cyclophosphamide was used in this study to define the contribution of the inflammatory response relative to direct cytopathic effects of guinea pig cytomegalovirus (GPCMV) in inducing sensorineural hearing loss in the guinea pig. The eighth nerve compound action potential (CAP) threshold on day 7 after inoculation of GPCMV into the scala tympani was an average of 35 dB greater for control animals than for those that were immunosuppressed with daily intraperitoneal injections of cyclophosphamide (20 mg/kg body weight). The amount of GPCMV antigen in the cochlea, detected immunohistochemically did not correlate with the CAP threshold. However, the greater the inflammatory response to GPCMV in the cochlea, the higher the CAP threshold and thus the greater the hearing loss. This study demonstrates that the inflammatory response to GPCMV may be more important than direct cytopathic effects of the virus in producing sensorineural hearing loss in GPCMV-induced labyrinthitis.

Electron Microscopic alterations in the fourth ventricle choroid plexus of rabbits after cyclophosphamide and mitomycin-C treatment

Cyclophosphamide and mitomycin-C have withstood the test of time for the treatment of many human neoplasms. While desirable results are achieved with their use, severe untoward toxic effects do occur in many body organs. Choroid plexus damage in the brain, varying from severe odema and necrosis of individual villi to haemorrhagic infarcts, are reported by Levine and Sowinski (1973) and Wenk et al (1979) after cyclophosphamide administration in rats. These authors noticed the choroid plex us of the fourth ventricle most susceptible for the drug injury. Separate groups of adult male and female rabbits were given single intraperitoneal injections of cyclophosphamide (200mg/kg) and mitomycin-C(0.1 mg/kg). After 48hrs, the control and treated animals were anaesthetized by using 30mg/kg sodium pentobarbital. The fourth ventricle choroid plexus was fixed in situ with buffered Karnovsky (1965) fixative (pH 7.3). After post-fixation in 1 per cent osmium tetroxide the tissues were processed for convential transmission electron microscopy.

Residual immunity of athymic NCr/sed nude mice and the xenotransplantation of human tumors

This study assessed the residual immunity possessed by NCr/Sed (nu/nu) athymic nude mice and examined strategies to reduce it in order to enhance the transplantability of human tumors for experimentation. Adult (8-week-old) female mice had fewer T cells (11%) and more B and NK (asialo-GM1-positive [ASGM1+]) cells in their spleen than euthymic (nu/+) controls. The number of phenotypically mature T cells increased with age, peaking at 16 weeks. ASGM1+ cells also increased in number over time, although the NK-activity decreased after 12 weeks. B cells remained relatively constant in number. Athymic NCr/Sed nude mice displayed reactivity against a human squamous carcinoma xenograph (FaDu), in a Winn's test and TD50 assay. Immunity against xenografts (TD50 assay) was significantly lower (by a factor of 2) in 4-week-old than in 12-week-old nude mice. Similarly, a significant 2-fold reduction in TD50 was obtained after a single intraperitoneal injection of cyclophosphamide into 8-week-old animals. Chronic (greater than 8 weeks) exposure of the nude mice to subcutaneously administered beta-estradiol markedly reduced the number of splenic NK cells and their cytolytic activity, but the TD50 reduction was not statistically significant (p = 0.1). Six Gray whole-body irradiations (WBI) had been shown to produce a highly significant, 3-fold reduction in the TD50 for FaDu. Flow cytometric analysis of splenic lymphoid cells from whole-body-irradiated recipients revealed: (a) marked initial depletion in the absolute numbers of lymphoid cells; (b) marked and long-lasting depletion of T cells, with slow and minimal recovery only evident between 6 and 12 weeks; (c) rapid, almost complete, depletion of B cells with prompt and partial recovery after 2 weeks; (d) depletion of NK cells and NK activity, with recovery by 10 weeks. No change in the number or phagocytic capacity of resident peritoneal macrophages was seen. These data give further support to a postulated role for residual T cells in the xenoreactivity of NCr/Sed nude mice.

Gastrointestinal and systemic candidosis in immunocompromised mice.

Oral-intragastric inoculation of 6-day-old outbred Crl:CFW(SW) BR mice with Candida albicans can lead to colonization of the gastrointestinal (GI) tract. We have shown that in the absence of an immunocompromising treatment, Candida is primarily localized in the stomach and intestines of mice at 20 days post-inoculation. Cultures of homogenates of the esophagus of most animals tested, and homogenates of the liver, lungs, spleen and kidneys of all animals tested, proved negative for C. albicans. Previous histological examinations of the GI tract of these colonized, non-immunocompromised mice showed hyphal elements associated with the stratified, squamous epithelium of the stomach in the region of the cardial-atrium fold. In this study, mice were immunocompromised by intraperitoneal injection of cyclophosphamide and cortisone acetate 11-14 days after oral-intragastric challenge with C. albicans and then sacrificed 20 days post-challenge. A high density of invasive hyphae was observed in the same, cardial-atrium region of the stomach of these animals. Cultures of the homogenized stomach showed a 100-fold increase in colony forming units (c.f.u.) of C. albicans compared with stomach homogenates of infected but non-immunocompromised controls. In addition, homogenates of the esophagus and selected body organs of most immunocompromised mice examined were positive for C. albicans by plate culture. When the immunocompromising drug treatment was delayed 3-5 weeks after oral-intragastric challenge, proliferation of C. albicans in the stomach and intestines was still evident, although fewer mice showed systemic spread and lower numbers of c.f.u. were recovered from body organ homogenates. Abscesses which contained both C. albicans hyphae and yeast cells were frequently observed in the liver and occasionally in the lungs and kidneys of immunocompromised mice sacrificed 20 days post-inoculation. The frequent occurrence of abscesses in the liver simulates a clinical variant of this mycosis, referred to as focal hepatic candidosis, which has been recognized with increasing frequency in immunocompromised patients. We suggest that the animal model described here may be particularly useful both for exploring methods which may prevent dissemination of C. albicans from localized foci of colonization in the GI tract after exposure of the host to immunocompromising drugs, and for testing the efficacy of anti-Candida drugs in clearance of the pathogen from body organs with established fungal abscesses.

顔面神経における単純ヘルペスウイルス感染の実験的研究

In the experimental animals, the author succeeded in establishing the pathogenicity of herpes simplex virus (HSV) in the facial nerve. KOS strain of HSV type 1 was inoculated to the tongue or the auricle of the balb/c mouse previously treated by intraperitoneal injection of cyclophosphamide. Between 4 and 12 days after the inoculation, the facial nerves were resected and submitted to histopathological and immunohistochemical investigations. Inflammatory changes were mainly observed in the geniculate ganglion and its proximal portion of the facial nerve irrespective of the site of the inoculation. In the same sites, HSV antigens were proved. The viral geniculate ganglionitis was induced in 21.4% of the animals inoculated to the tongue, 30.0% of those to the unilateral auricle, and 37.5% of those to the bilateral auricles respectively. This experiment demonstrated that HSV inoculated in the site innervated by sensory component of the facial nerve could affect the geniculate ganglion and its neighboring portion. These results presented a substantial support of the viral hypothesis in the etiology of Bell's palsy.

Expression of class II-MHC antigens by tumor-associated and peritoneal macrophages: systemic induction during tumor growth and tumor rejection.

It was shown previously that combination therapy of tumor-bearing mice resulted in amplification of antitumor responses at the site of tumor rejection and involved some of the classical components of the immunological network. As a continuation of this work, we now show that amplification of immune responses involves an increase in class II-MHC antigen (Ia) expression both at the tumor site and peripherally in the peritoneal cavity. Of further interest, however, was the observation that there was also an increase in Ia expression by tumor-associated macrophages (TAM) and peritoneal macrophages (PMs) in mice bearing progressing tumors. In these experiments, Ia expression by TAM and PMs was assessed in C57BL/6J mice bearing the progressing syngeneic MCA/76-9 sarcoma as well as in mice that had received combination therapy consisting of an intraperitoneal injection of cyclophosphamide (CY) and an intravenous injection of tumor-sensitized T lymphocytes (immune cells). When progressing tumors were analyzed, it was seen that by the fourth day after tumor cell implantation, 45-60% TAM expressed Ia, a level that was sustained throughout the course of tumor growth. The treatment of tumor-bearers with CY had no effect on Ia expression by TAM. However, the number of Ia-expressing TAM increased significantly after combination therapy and, as the tumors regressed, reached a peak of up to 100% in the second week after therapy. TAM were shown by Northern blot hybridization to contain mRNA encoding for the Ia beta chain. When Ia expression by PMs was assessed, it was seen that during tumor progression there was an increased expression of Ia over background (from less than 10 to about 40%), beginning 9-12 days after tumor cell implantation and continuing for the duration of the experiments. This was not influenced by CY injection. Combination therapy significantly increased the number of PMs expressing Ia (up to 80%). Ia expression by PMs could be induced rapidly in vivo by injecting normal B6 mice intraperitoneally (ip) with T cells isolated from tumors induced to regress by combination therapy. PMs isolated up to 15 days after injection were shown to express Ia. Moreover, the ip injection of a mixture of specific MCA/76-9 tumor cells and these tumor-associated lymphocytes resulted in a more rapid rate and a higher level of Ia expression by PMs compared with the level induced by either treatment alone.(ABSTRACT TRUNCATED AT 400 WORDS)

Correlation of natural killer cell activity and clearance of Cryptococcus neoformans from mice after adoptive transfer of splenic nylon wool-nonadherent cells.

Previous reports demonstrate that natural killer (NK) cells inhibit the growth of Cryptococcus neoformans in vitro, but conclusive evidence supporting the effectiveness of NK cells in host resistance to cryptococci is not available. The objective of these studies was to assess the ability of NK cells to clear C. neoformans from the lungs, livers, and spleens of infected mice. CBA/J mice were depleted of NK cells, as well as other natural effector cells, by an intraperitoneal injection of cyclophosphamide (Cy), 240 mg/kg of body weight. One day later, 7.5 X 10(7) nylon wool-nonadherent (NWN) spleen cells, either untreated or treated with anti-asialo GM1 and complement to remove NK cells, were adoptively transferred to Cy-pretreated mice. On day 2 after Cy treatment, the mice were injected intravenously with 2 X 10(4) cryptococci. At 4 and 6 days after Cy treatment, tissues were assayed for NK reactivity, using a 4-h 51Cr-release assay, and for in vivo clearance of cryptococci as reflected by mean log10 CFU per organ. We observed that Cy treatment depleted NK activity against YAC-1 targets and reduced in vivo clearance of C. neoformans from the tissues of infected mice. Additionally, Cy treatment depleted the total lung and spleen cellularity and the total number of peripheral blood lymphocytes when compared with those in normal untreated control mice. Also, spleen weights were significantly decreased in comparison with those of untreated animals 4 days after Cy treatment. Adoptive transfer of untreated NWN spleen cells into Cy-depressed mice restored the NK cell activity which correlated with enhanced clearance of cryptococci from lungs, livers, and spleens. In contrast, treatment of NWN spleen cells with anti-asialo GM1 and complement before adoptive transfer abrogated the ability of these cells to restore NK activity or reduce the numbers of cryptococci present in tissues of infected mice. Taken together, these data indicate that NK cells are the cells effective in diminishing the numbers of cryptococci in tissues of infected mice. Consequently, NK cells may play a role in first-line host resistance against C. neoformans.

The effect of cyclophosphamide on the allergic contact reaction in guinea pig: dose effects and influence on peripheral blood

Single intraperitoneal injections of cyclophosphamide were administered 6 days before testing guinea pigs sensitized to oxazolone in order to study the effects on inflammatory cell populations in blood and dermis. Skin tests were assessed macroscopically (erythema and oedema) and microscopically (counting of the dermal inflammatory cell infiltrate). At the highest dose (300 mg/kg) the allergic contact reaction was augmented with increases in erythema and oedema and the mononuclear dermal infiltrate. At the lowest dose (75 mg/kg), redness and oedema and all components of the dermal inflammatory cell infiltrate decreased. Total and differential white blood counts up to 20 days after administration of cyclophosphamide showed that a dose-dependent leukopenia maximal around 6 days occurred. During the leukopenia the differential count showed a lymphocytosis with a marked granulocyte depletion. The augmentation of the contact allergic reaction produced at the highest dose of cyclophosphamide occurs despite a marked peripheral blood leukopenia. Cyclophosphamide´s effects at the lower dose would appear to be of a non-specific anti-inflammatory nature.

Mutagenicity studies in somatic cells of mammals after inhalation and intraperitoneal injection of cyclophosphamide

Mutagenicity studies in somatic cells of mammals after inhalation and intraperitoneal injection of cyclophosphamide

Passive transfer of diabetes from BB/W to Wistar-Furth rats.

Autoimmune diabetes can be transferred to young, diabetes prone BB/W rats by injecting them intravenously with concanavalin A (Con A)-treated spleen cells from acute diabetic BB/W donors. This study describes the transfer of diabetes to the normal Wistar-Furth strain of rats using a similar procedure. For the successful transfer of diabetes it was necessary to immunosuppress recipient animals with a single intraperitoneal injection of cyclophosphamide 24-48 h before administering Con A-stimulated spleen cells from acute diabetic BB/W rats. Of 68 Wistar-Furth rats in immunosuppressed with a dose of 100-150 mg cyclophosphamide/kg body wt, 10 (15%) became diabetic. None of the control rats receiving either Con A-stimulated Wistar-Furth spleen cells (n = 28), freshly isolated BB/W spleen cells (n = 14), or fresh RPMI medium (n = 11) became diabetic. These data indicate that diabetes can be transferred from BB/W to Wistar-Furth rats. In addition, they support the hypothesis that cell-mediated immune processes are involved in the development of insulin-dependent diabetes and rule out any absolute requirement for BB-derived genes in the target pancreatic beta cells.

Immunohistochemical study of keratin in proliferative bladder epithelium induced by freezing, cyclophosphamide or N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide in the rat.

The distribution of intracellular keratin proteins was examined in a variety of urinary bladder lesions of the rat using the immunoperoxidase staining technique. In ethanol-fixed sections, the normal epithelium was strongly positive for keratin staining. Focal regenerative hyperplasia of the bladder epithelium induced by freezing exhibited relatively weak staining. However, diffuse regenerative hyperplasia induced by a single intraperitoneal injection of cyclophosphamide (CP) showed an intensely positive reaction throughout the epithelium. Of the sections fixed with Bouin's solution, the staining reaction was drastically reduced in the normal bladder and the staining was totally negative in the regenerative hyperplasia caused by freezing. Simple hyperplasia induced by a 4-week feeding of N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide (FANFT) as 0.2% of the diet exhibited strong reactivity, and nodular and papillarly hyperplasia induced by a 10-week feeding of FANFT was also positive for keratin throughout the lesions. In contrast to the preneoplastic lesions, FANFT-induced transitional cell carcinoma showed differential staining within the tumors. These results suggest that different keratin expression is involved in the proliferative bladder lesions induced mechanically by freezing and chemically by CP or by the carcinogen FANFT.