Reduction In Organ Weights Research Articles

Cardio-nephrotoxicity mediated by Echis ocellatus venom and its amelioration through kaempferol’s suppressive effect on oxidative stress, inflammation, and apoptosis expression

BackgroundEchis ocellatus venom toxins have the ability to impact multiple organ systems subsequent to envenomation. Kaempferol have been reported to have several therapeutic benefits. In this study, the therapeutic value of kaempferol was investigated in relation to the cardio-nephrotoxicity in rats resulting from E. ocellatus envenoming.MethodsFifty male wistar rats were allotted unbiased into five groups (n = 10) for this study. Group 1 was the control, while rats in groups 2 to 5 were envenomed with LD50 of E. ocellatus venom (0.22 mg/kg bw; i.p.). Group 2 was not treated after envenomation while groups 3, 4 and 5 were treated with polyvalent antivenom, 4 and 8 mg/kg of kaempferol, respectively.ResultsE. ocellatus envenomation caused considerable reduction in organ weight and relative organ weight in the envenomed untreated rats. The venom induced intense oxidative stress, inflammation, apoptotic damage to the cardiac and renal tissues accompanied with severe histomorphology in the organ tissues of untreated envenomed rats. In contrast, kaempferol treatment post-envenomation attenuated the venom-induced cardio-nephrotoxic responses in a dose dependent effect. Kaempferol substantially (p < 0.05) decreased malondialdehyde levels while enhancing reduced glutathione levels and superoxide dismutase and glutathione peroxidase activities in the heart and kidney of envenomed treated rats. Treatment of envenomed rats with kaempferol successfully decreased nitric oxide levels and myeloperoxidase activity. Overexpression of apoptotic caspase 3 and caspase 9 in cardiac and renal tissues were suppressed by kaempferol (p < 0.05). The histopathological result supports kaempferol’s ameliorative ability by convalescing the severe morphological alterations of cardiac and renal tissues induced by the venom.ConclusionFindings elucidate the significance of kaempferol as promising agent in the management of cardio-nephrotoxicity resulting from snakebite envenoming.

THU507 Tumor Activin A Mediates Cachexia Primarily Through White Adipose Tissue Hydrolysis In Pancreatic Ductal Adenocarcinoma

Abstract Disclosure: S. Yu: None. Y. Luan: None. R. Dong: None. A. Abazarikia: None. S. Kim: None. Pancreatic ductal adenocarcinoma (PDAC) commonly harbors mutations in KRAS and TP53 genes and its overall 5-year survival rate is only 10% in the US. Among risk factors associated with the dismal PDAC prognosis, cachexia accounts for about 30% of PDAC-related death and is present in nearly 80% of PDAC patients. Significant adipose tissue loss followed by skeletal muscle loss characterizes PDAC-related cachexia in humans and mice. Accumulated evidence indicates that activin A, a homodimer of inhibin βA that is encoded by the INHBA gene, is a recognized key cachexia mediator as supported by mitigation of weight loss by activin A antagonists in PDAC-bearing mice. However, much remains unclear regarding the primary source cells of activin A and the mechanisms by which activin A mediates cachexia in PDAC. To address the unknowns, we employed CRISPR/Cas9 system for Inhba gene knock-down (KD) in mouse KPC (KrasG12D and Trp53R172H) PDAC cells and generated orthotopic mice using Inhba KD or control KPC cells. Sham group which received the same operation except for cell injection was included. Body weight was measured on a daily basis, and blood and white adipose tissues (WATs) and skeletal muscle tissues were harvested on post-surgery day 12. We found that, when compared to the Sham group, injection of control KPC cells significantly elevated serum activin A levels up to 6 ng/mL and temporally decreased the average body weight in the orthotopic mice (Ctrl group), whereas Inhba gene KD significantly reduced serum activin A levels by ∼ 2 ng/mL and prevented weight loss in the orthotopic mice (KD group). When compared to the Sham and KD groups, notable reductions in organ weights of epididymal, mesenteric or subcutaneous WATs were found in the Ctrl group, however, weights for gastrocnemius and plantaris or soleus muscle tissues were statistically similar between the Ctrl and KD groups at post-surgery day 12. Correspondingly, the average serum free fatty acids levels were higher in the Ctrl group than the KD group. Histological observations support that dramatic adipocyte size reductions in WATs were prevented by serum activin A suppression. Immunofluorescence analysis showed that WATs from the Ctrl group had considerably higher protein expression of hormone-sensitive or adipose triglyceride lipase than the KD and Sham groups. Furthermore, phosphorylation of SMAD3 and phosphorylated PKA substrate which regulate adipocyte lipolysis were intensely stained in WATs from the Ctrl group when compared to the KD and Sham groups. Collectively, our findings identify PDAC cancer cells as the major source of activin A in PDAC and suggest systemic activin A as a potential mediator of PDAC-related adipose tissue loss. The current findings warrant further investigation to address clinical relevance. Presentation: Thursday, June 15, 2023

Ameliorative effects of palm oil on sniper induced toxicity on the cerebral cortex of adult male Wistar rats

Introduction: Sniper insecticide is a highly toxic, water soluble, synthetic organophosphate belonging to the DDVP (2, 2 — dichlorovinyl dimethylphosphate) compound family of chemicals. It’s wide spread indiscriminate indoor usage and more recently, an effective instrument for suicide related mortality which has been a source of great concern. Red palm oil, on the other hand, is an edible vegetable oil from the fleshy pulp of the oil palm that contains antioxidants; carotenoids that is responsible for its color (pigmentation) and Vitamin E that support brain health. This palm oil is consumed widely, used as an antidote in Nigeria and so is used for various ingested poisons. Aim: This study is aimed at investigating the ameliorative effects of palm oil on sniper induced toxicity on the cerebral cortex. Materials and methods: Sixteen adult male Wistar rats were randomly assigned to 4 groups (n = 4 per group). Group A was the Control, while Group B was administered just sniper (5 mg/kg), Group C was administered 2 ml of Red oil and Group D received 5 mg/kg of sniper + 2 ml of red oil. At the end of the experiment, the rats were sacrificed, their brains extracted and fixed in 10% formal saline for routine histological analysis. Results: There was significant increase in the body weight of Groups A, B, C and D. The relative organ weight of Wistar rats showed an insignificant decrease in the relative brain weight of Group B compared to the Control. Groups C and D also had a relative decrease in comparison to the control. The cerebral cortex reveals neuronal damage and proliferation of glial cells in the sniper-induced group and the mild recovery of neuronal cells in the rats treated with sniper and palm oil. Conclusion: The results from this study shows that there was a significant reduction in organ weight in Group B [Sniper only] when compared to the control group. When palm oil was administered, there was mild recovery of neuronal cells. This shows the ameliorative effects/ impact of Palm Oil in Neurotoxicity.

Methanolic Leaf Extract of Dryopteris dilatata Reverses Kidney Injury on Streptozotocin-Induced Diabetic Male Wistar Rats

Background: Hyperglycemia when sustained leads to diabetes which has become a chronic disorder having morbidity and mortality rate. This study investigated the effect of methanolic leaf extract of Dryoptersis dilatata (MEDd) on kidney injury caused on streptozotocin (STZ)-induced diabetic Wistar rats. Materials and Methods: Thirty Wistar rats were divided into five groups of six rats each. Group 1 received distilled water (10 ml/kg); Group 2 received STZ (60 mg/kg) only, Groups 3 and 4 received STZ followed by 400 and 800 mg/kg of MEDd, respectively, while Group 5 received STZ + Pioglitazone (10 mg/kg). After 2 weeks of treatment, rats were sacrificed and blood, spleen, liver, pancreas, and kidney were collected for biochemical analysis. Results: The results showed that MEDd extract caused a significant decrease (P &lt; 0.05) in STZ-induced diabetic rats, oxidative stress markers, malondialdehyde nitric oxide, and glutathione superoxide were ameliorated in organs such as the kidney and pancreas in diabetic rats after treatment with MEDd. Kidney markers (urea and creatinine) were ameliorated as well as reduction in organ weights in diabetic rats following treatment with MEDd. Conclusion: Therefore, it was observed from our study that MEDd has antidiabetic and nephron-protective capacity as it ameliorates in vivo adopted in lieu of nephropancreatic caused by STZ-induced diabetes.

BIOACCUMULATION OF HEAVY METALS AND HEALTH RISK ASSESSMENT AFTER THE CONSUMPTION OF SOME VEGETABLES GROWN ALONG RIVERS JAKARA AND GETSI IN ALBINO RATS

Cabbage and lettuce were collected from three locations each along the irrigation farms of Rivers Jakara and Getsi. Domestic and industrial waste are freely discharge into these rivers. Thirty Nine Albino Rats were divided into Thirteen (13) groups where Animals in groups 1-12 were fed with diet containing 10% cabbage or lettuce. The 13th group were fed with normal diet. After 90 days, the Animals were sacrificed, blood and organs were collected. Fe had the highest level in the blood and significantly different from the control in all the groups. The level of other essential heavy metals is in the following order of decreasing concentration, Fe&gt; Zn &gt; Cu &gt; Cr &gt; Mn &gt; Co while Pb has the highest concentration and in the following order of decreasing concentration Pb &gt; As &gt; Cd &gt; Ni &gt; Hg. A significant decrease in weight of the heart, kidney and pancreas was observed and a significance difference in the organ/body weight coefficient was observed in the Liver (group 3, 4 and 11) Kidney (group 5, 6 and 7) and Pancreas (group 1, 3 and 9). The bioaccumulation index (BAI) shows that eight (8), Seven (7) and Six (6) of the elements analyzed accumulates in the Liver and the Pancreas, Kidney and Heart respectively. Liver accumulated more Cadmium, Lead and Nickel while Pancreas accumulated Nickel, Manganese and Cobalt. Consumption of these vegetables led to reduction in organ weight, increase blood levels and bioaccumulation of some heavy metals in vital organs of albino rats

Triazine Derivative as Putative Candidate for the Reduction of Hormone-Positive Breast Tumor: In Silico, Pharmacological, and Toxicological Approach.

Background and objectives: Breast cancer is a heterogeneous disease that poses the highest incidence of morbidity among women and presents many treatment challenges. In search of novel breast cancer therapies, several triazine derivatives have been developed for their potential chemotherapeutic activity. This study aims to evaluate the N-nitroso-N-methyl urea (NMU)–induced anti–mammary gland tumor activity of 2,4,6 (O-nitrophenyl amino) 1,3,5-triazine (O-NPAT). Methods: The in silico modeling and in vitro cytotoxicity assay were performed to strengthen the research hypothesis. For in vivo experimentation, 30 female rats were divided into five groups. Group I (normal control) received normal saline. Group II (disease control) received NMU (50 mg/kg). Group III (standard control) was treated with tamoxifen (5 mg/kg). Groups IV and V received O-NPAT at a dose level of 30 and 60 mg/kg, respectively. For tumor induction, 3 intraperitoneal doses of NMU were given at a 3-week interval, whereas all treatment compounds were administered orally for 14 consecutive days. Biochemical and oxidative stress markers were estimated for all experimental animals. DNA strand breakage alongside inflammatory markers was also measured for the analysis of inflammation. The hormonal profile of progesterone and estrogen was also estimated. Results: The test compound presented a significant reduction in organ weight and restored the hepatic and renal enzymes. O-NPAT treatments enhanced the antioxidant enzyme level of catalase (CAT), superoxide dismutase (SOD), and total sulfhydryl (TSH), with a highly significant reduction in lactate dehydrogenase (LDH) and lipid peroxidation. Also, the decrease in fragmented DNA, hormonal levels (estradiol and progesterone), and inflammatory cytokines (IL-6 and TNF-α) justified the dosage efficacy further supported by histopathological findings. Conclusion: All results indicated the anti–breast tumor activity of O-NPAT and presented its possibility of exploitation for beneficial effects in breast cancer treatment.

Aspartame induces alteration in electrolytes homeostasis of immune organs in wistar albino rats

Aspartame is rapidly and completely metabolized in humans and experimental animals to aspartic acid (40%), phenylalanine (50%) and methanol (10%). Methanol, a toxic metabolite is primarily metabolized by oxidation to formaldehyde and then to formate. These processes are accompanied by the formation of superoxide anion and hydrogen peroxide. This study is focused to understand whether the oral administration of Aspartame (40mg/kg bw) for 15 days, 30 days, and 90 days have any effect on immune organs. Damage to plasma membrane was assessed by levels of membrane-bound ATPases. Oxidative stress status was assessed by alterations in level of lipid peroxides, protein carbonyls, protein thiol and lipid-soluble antioxidant vitamin E. To mimic human methanol metabolism, folate-deficient animals were used. There was decrease in all membrane-bound ATPases activities in immune organs. Aspartame administration to rats inducing excess free radical generation is confirmed by increase in lipid peroxidation, obvious which is also again substantiated by the elevated protein carbonyl and decrease in protein thiol in this study. These excess free radical generations also decrease the cellularity (reduction in organ weight and cell count) of immune organs.

Effect of aspartame on lipid peroxidation and membrane bound enzymes in immune organs of rats

Objective: The artificial dipeptide sweetener aspartame (L-aspartyl-L-phenylalanine methyl ester) is present in many products especially unsweetened and sugar products. These products are frequently utilized by people trying to lose weight or patients with diabetes. Concern relating to the possible adverse effect has been raised due to aspartame’s metabolic components. Aspartame is rapidly and completely metabolized in humans and experimental animals to aspartic acid (40%), phenylalanine (50%) and methanol (10%). Methanol, a toxic metabolite, is primarily, metabolized by oxidation to formaldehyde and then to formate: these processes are accompanied by the formation of superoxide anion and hydrogen peroxide. Methods: This study focus is to understand whether the oral administration of aspartame (40 mg/kg body weight) for 90 days has any effect on lipid peroxidation, nitric oxide level, membrane bound ATPases of immune organs and differential leucocyte count of rats. Results: After 90 days of aspartame administration, there was a neutrophil and lymphocyte imbalance in normal white blood cell homeostasis, a significant increase in lipid peroxidation with nitric oxide level, and an alteration of membrane bound ATPase activities, which finally decreased the cellularity (reduction in organ weight and cell count) of immune organs. Conclusion: Aspartame metabolite methanol or formaldehyde may be the causative factors behind the changes observed. This study concludes that oral administration of aspartame for longer duration may cause oxidative stress on immune organs.

Oxidative stress in the blood of pregnant rats and reduction of organ weights caused by the polychlorinated biphenyl congener 77

3,3′,4,4′-Tetrachlorobiphenyl (PCB 77), one of the environmentally persistent polychlorinated biphenyls that have been used for industrial purposes, was repeatedly administered to pregnant rats by gavage at a dose of minimal, non-lethal toxicity in order to study its effects on routine blood variables, parameters of oxidative stress and hematopoietic organs in pregnant female rats. Of the routine blood parameters, PCB 77 reduced the mean corpuscular hemoglobin concentration and platelet counts; the levels of malondialdehyde and glutathione peroxidase in the blood were higher during the first days of gestation (1–7), as compared to the respective controls. In the subsequent period between days 8 and 18, these parameters did not show any significant change after PCB 77 treatment. Routine blood parameters and oxidative stress parameters indicated a moderate degree of oxidative stress which alone could not bring about the serious weight reduction of the thymus, pituitary, and kidneys. The extent of oxidative stress did not correlate with the weight reduction of some of the blood-forming organs.

NS-018, a Potent Novel JAK2 Inhibitor, Effectively Treats Murine MPN Induced by the Janus Kinase 2 (JAK2) V617F Mutant

AbstractAbstract 4106An activating mutation in the Janus kinase 2 gene (JAK2) (G1849T, which produces JAK2 V617F) occurs at a high frequency in Bcr-Abl-negative myeloproliferative neoplasms (MPNs). JAK2 V617F induces cytokine-independent growth in cell lines and, in murine models, recapitulates much of the pathobiology observed in MPN patients, suggesting that small-molecule inhibitors targeting JAK2 may be therapeutically useful. Some orally bioavailable inhibitors of JAK2 are already in clinical trials.NS-018 is a novel JAK2 inhibitor that inhibits JAK2 enzyme activity with an IC50 value of less than 1 nM. NS-018 shows 30–50-fold selectivity for JAK2 over other JAK-family kinases such as JAK1, JAK3 and TYK2.We tested NS-018 in a murine model of MPN induced by JAK2 V617F. Mice expressing JAK2 V617F controlled by the H2Kb promoter (V617F-TG mice) show an MPN phenotype: leukocytosis, thrombocytosis, progressive anemia, hepatosplenomegaly with extramedullary hematopoiesis, megakaryocyte hyperplasia and bone marrow fibrosis. They also exhibit body weight loss and high mortality compared to wild-type controls. Bone-marrow cells show constitutive activation of STAT5 and cytokine-independent growth of erythroid colony-forming units (CFU-E).NS-018 inhibited cytokine-independent CFU-E growth and constitutive activation of STAT5 in V617F-TG cells in vitro. For in vivo experiments, V617F-TG mice were divided into treatment and vehicle control groups after disease was established at 12 weeks after birth. NS-018 was administered for 24 weeks by oral gavage at doses of 25 mg/kg or 50 mg/kg bid, and the control groups received vehicle only. Mice were monitored by blood counts, and a subset of mice was euthanized for detailed histopathology and fluorescence activated cell sorting analysis. During the study, 12 of 34 mice died in the vehicle group, whereas 1 of 36 mice died in the 50 mg/kg group. There was a statistically significant prolongation of survival in the 50 mg/kg group (p<0.01). Mice treated with NS-018 gained more weight than vehicle-treated mice, and were comparable to wild-type mice. V617F-TG at 12 weeks old showed severe leukocytosis with average white blood cell counts of 24 × 1010/L. After two weeks of NS-018 treatment, the leukocyte count was reduced to 59% in 25 mg/kg group and 39% in the 50 mg/kg group compared to vehicle group, and the effect was maintained until the end of the study. The inhibitory effect of NS-018 on T or B lymphocytes was much less than on myeloid cells. The 50 mg/kg group showed no progression of anemia. NS-018 treatment also improved hepatosplenomegaly in a dose-dependent manner. In the spleen, Mac-1/Gr-1+ myeloid cells associated with extramedullary hematopoiesis were significantly decreased, and B220+ B cells were increased by NS-018 treatment. In correlation with reduction of organ weights and infiltrating myeloid cells, there was also clear evidence of a dose-dependent reduction in the histopathology of extramedullary hematopoiesis in the spleen, liver, and lungs of NS-018-treated mice. In contrast to the improvement in the pathology of these organs, NS-018 had little impact on the progression of fibrosis and megakaryocyte hyperplasia in bone marrow. No significant toxicity was observed in treated mice.In conclusion, NS-018 demonstrated therapeutic efficacy in a murine model of MPN induced by JAK2 V617F. In V617F-TG, which closely mimics human MPN, NS-018 significantly improved survival, body weight loss, hepatosplenomegaly, leukocytosis and anemia progression, thus confirming the viability of a targeted-therapy approach in managing JAK2 V617F positive MPNs. On the basis of these preclinical experiments, NS-018 appears to be an excellent candidate for phase I/II studies in patients with Bcr-Abl-negative MPNs. Disclosures:Nakaya:Nippon Shinyaku Co., Ltd: Employment. Homan:Nippon Shinyaku Co., Ltd: Employment. Kotera:Nippon Shinyaku Co., Ltd: Employment. Shibayama:Nippon Shinyaku Co., Ltd: Employment. Naito:Nippon Shinyaku Co., Ltd: Employment. Shimoda:Nippon Shinyaku Co., Ltd: Research Funding.

Standard organ weights among elderly Japanese who died in hospital, including 50 centenarians

The present study was conducted to determine standard organ weights among the elderly, because little has been reported on this subject. To analyze the human aging process in terms of organ weights, age-associated changes and correlations among organ weights and the contributions of age and nutrition to organ weights were also examined. The subjects included 1615 consecutive autopsy cases of patients aged 60-99 years who died between 1995 and 2003, and cases of 50 centenarians who died between 1973 and 2005. The weights of nine organs were measured before formalin fixation. If affected by serious diseases, such as cancer, the organs were excluded from the study. Values beyond 99% of the bilateral measurement limits were also excluded. In this manner the standard organ weights were obtained. The organ weights decreased significantly according to age in all organs except for the heart in men and the lungs in both genders. Undernutrition strongly contributed to organ weight except for the hypophysis, and was especially apparent in the heart and liver. In conclusion, the standard organ weights of elderly patients who died while hospitalized were determined. Undernutrition contributed significantly to a reduction in organ weights.

Calcineurin A-α But Not A-β Is Required for Normal Kidney Development and Function

Calcineurin is an important signaling molecule in the kidney and may be involved in a variety of processes. The phosphatase subunit of calcineurin (CnA) has three isoforms, alpha, beta, and gamma. In this study, we investigated the effect of loss of calcineurin A-alpha (CnA-alpha) or calcineurin A-beta (CnA-beta) on the development and function of the kidney. Total calcineurin expression and activity was significantly reduced in whole kidney homogenates from both CnA-alpha -/- and CnA-beta -/- mice. Kidneys of CnA-beta -/- mice appear normal and the mice develop with no phenotypic abnormalities. In contrast, kidneys of CnA-alpha -/- animals fail to fully develop. In particular, postnatal maturation of the nephrogenic zone (NZ) is defective. Within the NZ, glomeruli also fail to mature and lack mesangial cells. In addition to alterations in development, there is an absence of proliferation and an increase of cell death in the NZ with loss of CnA-alpha. Finally, increased collagen deposition is observed and serum creatinine levels are significantly increased in CnA-alpha -/- animals compared to wild-type littermates, indicating that kidney function is impaired. In summary, absence of CnA-alpha but not CnA-beta leads to a defect in normal maturation of the NZ and glomeruli, alterations in the cell cycle, and impaired kidney function.

Amelioration of doxorubicin-induced cardiac and renal toxicity by pirfenidone in rats.

Doxorubicin (DXR) is an anthracycline glycoside with a broad spectrum of therapeutic activity against various tumors. However, the clinical use of DXR has been limited by its undesirable systemic toxicity, especially in the heart and kidney. This study was designed to test the effectiveness of dietary intake of pirfenidone (PD) against DXR-induced cardiac and renal toxicity. Male Sprague Dawley rats were placed into four treatment groups: saline injected intraperitoneally (i.p.) plus regular diet (SA+RD); DXR i.p. plus regular diet (DXR+RD); saline i.p. plus the same diet mixed with 0.6% PD (SA+PD); and DXR i.p. plus the same diet mixed with 0.6% PD (DXR+PD). The animals were fed regular or regular plus PD diets 3 days prior to i.p. injections of either saline or DXR and continuing throughout the study. A total dose of DXR (16.25 mg/kg) or an equivalent volume of saline was administered in seven injections (2.32 mg/kg per injection) three times per week with an additional dose on the 12th day. At 25 days following the last DXR or saline injection, some animals were anesthetized for the measurement of cardiac and pulmonary function, and others were killed by an overdose of pentobarbital. At the time the animals were killed, abdominal fluid was collected. Kidney and heart were removed, weighed, fixed with 10% formalin or frozen in liquid nitrogen. The fixed tissues were used for histological examination and the frozen tissues were used for biochemical studies. The average volumes of abdominal fluid in the DXR+RD and DXR+PD groups were 9.42 ml and 3.42 ml and the protein contents of abdominal fluid in the DXR+RD and DXR+PD groups were 218 mg and 70 mg, respectively. A 12.5% mortality occurred in the DXR+RD group as compared to 0% in DXR+PD group. There were no changes in any of the cardiac or pulmonary physiological parameters in any of the four groups. The changes in the heart and kidney of the DXR+RD group included reduction in organ weight, increase in hydroxyproline content of heart, increase in hydroxyproline, and lipid peroxidation in the kidney and plasma, and increase in protein concentration in urine as compared to rats in the control, SA+RD and SA+PD groups. Treatment with PD abrogated the DXR-induced increases in hydroxyproline content in the heart and kidney, lipid peroxidation of the kidney and plasma, and protein content of the urine in the DXR+PD group. DXR treatment alone caused disorganization of cardiac myofibrils, vacuolization of the myofibers, and renal tubular dilation with protein casts in both the cortical and medullary regions. Treatment with PD minimized the DXR-induced histopathological changes of heart and kidney in the DXR+PD group. Treatment with PD reduced the severity of DXR-induced toxicity as assessed by reduced mortality, diminished volume of recovered fluid in the abdominal cavity, and severity of cardiac and renal lesions at both the biochemical and morphological levels. These results indicate that PD has the potential to prevent DXR-induced cardiac and renal damage in humans on DXR therapy.

Oral (gavage) two-generation (one litter per generation) reproduction study of pentachlorophenol (penta) in rats.

The potential for pentachlorophenol (penta) to induce general and reproductive/developmental toxicity was evaluated in Crl Sprague-Dawley rats, employing a two-generation reproduction toxicity study. Penta was administered by gavage at doses of 0, 10, 30, and 60 mg/kg/day. In both generations, the parental animals (30/sex/group) were intubated daily for 10 weeks before cohabitation and continuing through cohabitation, gestation, and lactation periods. Intubation of the F1 generation was begun 28 days postpartum. Animals were evaluated daily for mortality and general toxicity (clinical observations, body weights and gains, feed consumption). Organ weights were recorded and histopathological evaluations were made. Specific indices of reproductive function evaluated included estrous cycles, mating and fertility, parturition, lactation, viability, and growth and development of offspring, including sexual maturation, sperm parameters, and numbers of ovarian primordial follicles. All deaths in the parental rats were unrelated to penta. Expected metabolic effects of penta, sporadic increased liver weights associated with hepatocellular centrilobular hypertrophy and vacuolation and lipofuscin pigmentation, were evident in the 10-, 30-, and 60-mg/kg/day dose group P1 and F1 animals. Toxicity, in the form of liver pathology (single cell necrosis), reduced body weights and associated reductions in organ weights, and reduced feed consumption were noted in both generations at the 30- and 60-mg/kg/day doses. Developmental toxicity associated with these doses included reduced pup weights and viability. The 60-mg/kg/day dose also resulted in delayed sexual maturation, decreased spermatid counts, small prostates and testes, decreased implantations, reduced fertility, and increased resorptions of embryos. Based on these results, it was concluded that 30 mg/kg/day is the lowest-observable-adverse-effect level (LOAEL) and 10 mg/kg/day is the no-observable-adverse-effect level (NOAEL) for both reproductive and general toxicity. These findings are consistent with results from previously conducted studies wherein reproductive/developmental toxicity was observed only at doses that also induced general toxicity. It differs from previous findings in that the NOAEL for general toxicity is two to three times higher for the more pure product than for products produced and tested previously. In addition, the results did not indicate bioaccumulation of penta. Thus, penta did not selectively affect reproduction or development of the offspring of rats at a dose of 10 mg/kg/day, a dose that is 7000 to 20,000 times higher than human exposure.

Bacille Calmette-Guérin (BCG)-associated inflammation and fibrosis: modulation by recombinant BCG expressing interferon-gamma (IFN-gamma).

Immunization with existing BCG vaccines has failed to confer consistent protection against tuberculosis. One of the ways to improve the efficacy of BCG is by enhancing its ability to induce a type-1 T cell response. However, this approach carries the risk that enhanced immunoreactivity may exacerbate tissue pathology associated with vaccination. The aim of the present study was to determine whether use of a recombinant BCG expressing IFN-gamma (BCG-IFN) would result in an alteration in the pattern of inflammation and local tissue fibrosis. A murine intravenous BCG infection model was used in which there was a time- and dose-dependent increase in the weight and number of granulomas in the liver. Infection was associated with increased inflammatory activity in the liver, as shown by the increase in expression of inducible nitric oxide synthase (iNOS) assessed by immunochemistry and by measurement of specific mRNA, and in fibrosis measured by hydroxyproline content of the liver and percentage of granuloma cells staining positively for type 1 procollagen. Infection with BCG-IFN resulted in a reduction in organ weight and bacterial load on day 21 compared with infection with control BCG transformed with vector alone (BCG-plasmid). By day 21, there was also a reduction in iNOS mRNA and iNOS+ cells in granulomas in mice infected with BCG-IFN compared with infection with BCG-plasmid, and a similar reduction in both total number of granulomas and liver hydroxyproline content. These results demonstrate that the granulomas in the areas of mycobacterial infection are active sites of both inflammation and fibrosis, and that the local expression of IFN-gamma by the recombinant BCG results in more efficient bacterial clearance which is accompanied by a reduction in tissue pathology.

Evaluation of the potential immunotoxicity of chlorinated drinking water in mice

Recent epidemiological studies have reported associations between the consumption of chlorinated drinking water and various types of human cancer; in addition, exposure to chlorine (Cl −) in drinking water has been reported to suppress certain immune functions in laboratory animals. The current studies were conducted to extend our knowledge of the effects of drinking water exposure to Cl −. Female C57BL/6 mice were administered hyperchlorinated drinking water (7.5, 15, or 30 ppm Cl −) for 2 weeks prior to sacrifice for evaluation of spleen and thymus weights, the plaque-forming cell (PFC) response, hemagglutination (HA) titer, and lymphocyte proliferation (LP). Significant reductions in organ weights and immune response were observed in the positive control groups (i.e. dexamethasone- or cyclophosphamide-exposed mice). No consistent differences were observed between the Cl −-exposed animals and vehicle control mice for the evaluated parameters. Thus, under the conditions of these experiments, 2 weeks of exposure to hyperchlorinated drinking water had no apparent adverse effects on immune function.

In utero and lactational exposure of male rats to 2,3,7,8-tetrachlorodibenzo- p-dioxin : I. Effects on androgenic status

When administered in overtly toxic doses to postpubescent rats, 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) produces a variety of adverse effects on the male reproductive system including a decrease in plasma androgen concentrations. If such an androgenic deficiency were produced prenatally and/or early postanatally it could potentially impair male reproductive function by disrupting the development of sex organs and/or causing incomplete sexual differentiation of the central nervous system. To determine whether TCDD can reduce androgen concentrations perinatally and/or impair androgen-dependent perinatal development, pregnant Holtzman rats were treated with 1.0 μg TCDD/kg or vehicle on Day 15 of gestation. Plasma testosterone concentrations in fetal males were significantly reduced by TCDD on Gestation Days 18 through 21. The surge in plasma testosterone concentrations shortly after birth was also significantly reduced, as was anogenital distance, an androgen-dependent parameter. To further investigate the effects of perinatal TCDD exposure on the male reproductive system, rats born to dams given TCDD (0.064, 0.16, 0.40, or 1.0 μ/kg, po) or vehicle on Day 15 of gestation were evaluated from birth through sexual maturation. This report describes their growth, physical development, and androgenic status (i.e., androgen concentrations and androgen-dependent structures and functions); effects on spermatogenesis, testicular histology, sexual behavior, and fertility are reported separately. There was little evidence that TCDD caused maternal toxicity. Signs of overt toxicity in offspring were limited to an 8% reduction in live births (highest dose only) and to decreases in body weight gain and feed consumption (two highest doses only) which disappeared by early adulthood. With respect to androgenic status, maternal TCDD doses as low as 0.16 μg/kg produced significant dose-related decreases in the anogenital distance of 1- and 4-day-old males, delays in testicular descent, and decreases in seminal vesicle and ventral prostate weights. The reductions in organ weights were observed when rats were at the juvenile, pubertal, postpubertal, and mature stages of sexual development. Plasma testosterone and 5α-dihydrotestosterone concentrations tended to be reduced at these times (though not significantly), while plasma luteinizing hormone concentrations were generally unaffected. Collectively, these results demonstrate that perinatal TCDD exposure alters the androgenic status of male rats from the fetal stage into adulthood, and that TCDD can affect androgenic status without causing overt toxicity. In rats, the male reproductive system appears to be more sensitive to the toxic effects of in utero and lactational TCDD exposure than any other organ or organ system studied thus far.

Effects of Fetal Insulin Deficiency on Growth in Fetal Lambs

Insulin may be an important regulator of growth in late fetal life. To assess the importance of endogenous insulin release in regulation of normal fetal growth, eight fetal lamb pairs were given either an intravenous injection of streptozocin (STZ), a nitrosourea that selectively damages pancreatic beta-cells, or buffer infusion (controls). In six preparations, twins were used, and in two cases, triplets, thus allowing for comparison between treated and control fetuses residing in the same intrauterine environment. Fetal STZ injection was associated with relative fetal hyperglycemia, hypoinsulinemia, and a decrease in the fetal plasma insulin-glucose ratio. Fetal lambs exposed to STZ also developed a mild nonprogressive metabolic acidosis compared with controls. Fetal body weight was depressed by 21% overall, the magnitude of reduction related to length of time in utero after STZ injection. Similar reductions in organ weights (liver, heart, and kidney) were also observed in STZ-administered fetuses compared with controls. Protein accretion in carcass, liver, and kidney after STZ was also depressed, but no significant changes in fetal lipid accretion were observed. Skeletal growth, as measured by tail and tibial lengths, was also depressed after STZ but to a lesser extent than body weight or protein accretion. Thus, in a stable maternal environment, isolated fetal insulin deficiency is associated with significant retardation of somatic and skeletal growth and protein deposition.

Preliminary Toxicity Data on New Ethylenediamine Derivatives Designed for Iron Chelation

During development of iron chelators for the treatment of iron overload in Cooley's anemia, the opportunity arose to obtain some acute and subacute toxicity data on hydroxybenzyl-ethylenediamine (HBED) derivatives, hydroxyphenyl-ethylenediamine (HPED) congeners, and ethylenediamine-phenylacetic acid (EDHPA) compounds. Such data permitted correlations among structure, toxicity, and efficacy parameters useful for further syntheses. Young, adult male mice were used to perform acute intraperitoneal (IP) or oral (PO) LD50 studies, and hypertransfused groups were treated IP or PO with vehicle, Desferal, or test chelators once daily for 7 days. Changes in behavior, weights of spleen, liver, and feces, and urine volume and gross pathology were monitored. In the HBED series, esterification of acetic acid groups with or without acetylation of phenolic hydroxyl groups increased LD50 values, and the major behavioral change was decreased exploratory activity. Dimethylenephosphonate derivatives of HBED were more toxic. Single treatment with compounds in the EDHPA and HPED series yielded IP LD50 values between 175 and 325 mg/kg. After subacute treatment, active compounds were found in each series, and efficacy was related to toxicity. Chelation effectiveness was usually greater by the IP route than orally. The major toxic signs were reduction in organ weights and excreta output.

Effect of somatostatin on the growth of gastrointestinal mucosa and pancreas in rats. Role of endogenous gastrin.

This study was undertaken to determine the influence of somatostatin on the growth of gastric, duodenal, and pancreatic tissue in rats placed on liquid diet. In the first part of the study animals were fed an elemental liquid diet for 10 days and then killed, and the growth of the oxyntic gland area of the stomach, 2 cm segments of duodenum and pancreatic tissue was determined. Feeding an elemental diet caused a decrease in organ weight, nucleic acid contents and serum gastrin level. Subsequent addition of pentagastrin prevented this reduction in organ weight and RNA and DNA contents while somatostatin failed to affect the decrease in growth parameters or serum gastrin level in tests with or without addition of pentagastrin. In a second part of the study, sham operated and antrectomised rats were used. Antrectomy caused a significant decrease in serum gastrin concentration and resulted in a significant reduction in the weight and RNA and DNA contents of the tissue tested. As in liquid diet, subsequent administration of pentagastrin prevented the reduction in the growth parameters both in tests with and without somatostatin. These results suggest that somatostatin inhibits the growth of the gastroduodenal mucosa by a mechanism involving, at least in part, the suppression of gastrin release.