(2-Carboxyethyl) dimethylsulfonium bromide supplementation in non-fish meal diets for on-growing grass carp (Ctenopharyngodon idella): Beneficial effects on immune function of the immune organs via modulation of NF-κB and TOR signalling pathway.

The immune function of immune organs is extremely crucial for maintaining organism health status, which ultimately affects fish growth. Our previous study has found that dietary supplementation of (2-carboxyethyl)dimethylsulfonium Bromide (Br-DMPT) in non-fish meal (NFM) diet could promote the growth of grass carp (Ctenopharyngodon idella), whereas the underlying reason or mechanism for this results is largely unclear. Herein, we further explored whether dietary supplementation of Br-DMPT promoted fish growth is connected with the enhanced immune function in the immune organs (the head kidney, spleen and skin). In this study, 540 fish (216.49 ± 0.29 g) were irregularly distributed to six groups with three replicates (30 fish replicate-1) and fed corresponding diets group containing a fish meal (FM) diet group and five different NFM diets supplemented with gradational Br-DMPT (0-520.0 mg/kg level) group for 60 days. After the 60-days feeding trial, 8 fish from each replicate were selected and then conducted a challenge test with A. hydrophila for 14 days. Our results indicated that in the NFM diets, appropriate Br-DMPT: (1) significantly decreased the morbidity of skin haemorrhage and lesion after A. hydrophila infection (P < 0.05). (2) significantly improved the innate and adaptive immune components (lysozyme, complement 3, immunoglobulin M and antibacterial peptides et al.) (P < 0.05). (3) increased the gene expressions of main anti-inflammatory cytokines partially by referring to TOR signalling pathway, and decreased the gene expressions of main pro-inflammatory cytokines partially by referring to NF-kB signalling pathway (P < 0.05). Strikingly, no statistical difference could be found in the most of above immune parameters between 260.0 mg/kg Br-DMPT diet group and FM diet group (P > 0.05). Taken together, in non-fish meal diet, appropriate supplementation of Br-DMPT could improve the disease resistance capacity, non-specific immunity and ameliorate inflammation, and simultaneously could mitigate these adverse effects induced by the non-fish meal diet in fish immune organs. Moreover, this study may be helpful to decipher the underlying mechanisms of how Br-DMPT promote fish growth by immune organs and also provide scientific theoretical evidence for the future application of Br-DMPT as a new immunopotentiator in aquaculture industry.

Protective effects and potential mechanisms of (2-Carboxyethyl) dimethylsulfonium Bromide (Br-DMPT) on gill health status of on-growing grass carp (Ctenopharyngodon idella) after infection with Flavobacterium columnare.

In this study, the protective effects and potential mechanisms of (2-Carboxyethyl) dimethylsulfonium Bromide (Br-DMPT) were evaluated in relation to the gill health status of on-growing young grass carp (Ctenopharyngodon idella). A total of 450 grass carp (216.49 ± 0.29 g) were randomly distributed into five treatments of three replicates each (30 fish per replicate) and were fed diets supplemented with gradational Br-DMPT (0-520.0 mg/kg levels) for 60 days. Subsequently, the fish were challenged with Flavobacterium columnare for 3 days, and the gills were sampled to evaluate antioxidant status and immune responses evaluation. Our results showed that, when compared to the control group, dietary supplementation with appropriate Br-DMPT levels resulted in the following: (1) decreased gill rot morbidity and improved gill histological symptoms after exposure to F. columnare (P < 0.05); (2) improved activities and gene expression levels (except GSTP2 gene) of antioxidant enzymes and decreased oxidative damage parameter values (reactive oxygen species, malondialdehyde and protein carbonyl) (P < 0.05), which may be partially associated with the nuclear factor-erythroid 2-related factor 2 (Nrf2) signalling pathway (P < 0.05); (3) increased lysozyme (LZ) and acid phosphatase (ACP) activities and complement 3 (C3), C4 and immunoglobulin M (IgM) contents, and upregulated genes expressions of antibacterial peptides (liver-expressed antimicrobial peptide-2A, -2B, hepcidin, ß-defensin and mucin2) (P < 0.05); (4) upregulated gene expressions of anti-inflammatory cytokines (except IL--4/13B) that may be partially to the TOR/(S6K1, 4E-BP1) signalling pathway, and downregulated gene expressions of pro-inflammatory cytokines (except IL-12P35) may be partially to the IKK ß, γ/IκBα/NF-kB) signalling pathway (P < 0.05). Taken together, our results indicate that dietary supplementation with appropriate amounts of Br-DMPT may effectively protect on-growing grass carp from F. columnare by strengthening gill antioxidant capacity and immunity. Furthermore, based on measures of combatting gill rot, antioxidant indices (MDA) and immune indices (LZ), the dietary Br-DMPT supplementation levels for on-growing grass carp are recommended to be 291.14, 303.38 and 312.01 mg/kg diet, respectively.

Hyperhomocysteinemia can be ameliorated by dimethylsulfoniopropionate in place of folic acid in mice.

Acute homocysteinemia mice were prepared by forcibly oral administration of homocysteine (4 mM, 2 mL). The amounts of plasma homocysteine were estimated by a fluorescence method with HPLC. Folic acid (0.6 mM, 2 mL), DMSP, or betaine (20 mM, 2 mL each) was intraperitoneally administrated into the mice suffered from the acute homocysteinemia on the 20th, 40th or 60th min after the oral supplementation of homocysteine, then amounts of plasma homocysteine were determined by the HPLC method 40 min after each addition, respectively. The results indicated that the intraperitoneal addition of folic acid or DMSP in this order of the 40th, 60th and 20th min after the oral supplementation of homocysteine significantly reduce the quantities of plasma homocysteine, but betaine exerted the fairly lesser effects. The amounts of homocysteine without any additive linearly and rapidly appeared to increase up to 60 min, at which those were about 8-12 fold the normal levels of homocysteine, and thereafter decreased in these experiments. Accordingly, folic acid which is known to effectively improve homocysteinemia was proven to be completely replaced by DMSP under the experimental conditions.

Direct effects of high concentrations of dimethylsulfoniopropionate, vitamin E and ferulic acid on the senility of aged scenescene-accelerated mouse (SAMP8).

The effects of high concentrations of dimethylsulfoniopropionate (DMSP), vitamin E and ferulic acid solutions on the aging of male and female scenescene-accelerated mouse (SAMP8) at the age of 50 wk were examined by direct supplementation to their stomachs twice a week for 28-30 d. The addition of 1 mL of DMSP, vitamin E and ferulic acid solutions (21% each) to the male and female mice in each group in this order rather elevated their growth and significantly suppressed their total grading score and loss of learning and memory with increasing rearing times for the short experimental period. However, there were no significant differences found between the male and female mice during the experimental period. The antioxidant and hormonal actions for the effects of the test compounds on the aged SAMP8 were possibly considered.

[A fifty two-week oral repeated dose toxicity study of suplatast tosilate (IPD-1151T) in dogs].

A 52-week oral repeated dose toxicity study of suplatast tosilate (IPD-1151T), a newly developed anti-allergic agent, was carried out in beagles by oral administration of 30, 90, 270 and 810 mg/kg/day for 52 weeks. The recovery study was carried out by the withdrawal for 5 weeks using control and the 810 mg/kg groups. The results are as follows: 1. Observation of general conditions revealed soft feces, mucous feces, and diarrhea in both sexes of the 270 and 810 mg/kg groups during the administration period, and these findings disappeared during the withdrawal period. One female of the 810 mg/kg group exhibited tremors in the legs and neck, staggering, a decrease of spontaneous motor activity, and clonic convulsions in Week 17 of administration and died on Day 118. One male of the same group exhibited whole body tremors and staggering from Week 32 to Week 52. 2. Body weight gain tended to be inhibited in males of the 810 mg/kg group during the administration period. The body weight of the female that died decreased rapidly after the appearance of neurological symptoms. The body weight of the male that exhibited neurological symptoms decreased after their appearance but later increased. 3. There were no abnormal changes in food consumption in all of the sacrificed dogs. The female that died did not eat at all after the appearance of neurological symptoms. The male that exhibited neurological symptoms did not eat at all for 1 week after their appearance, but the food consumption returned to normal thereafter. 4. Prothrombin times were prolonged in males of the 270 and 810 mg/kg groups at Week 26, and activated partial thromboplastin times were prolonged in males of the 810 mg/kg group at Week 52. 5. Plasma levels of alkaline phosphatase, GPT and LDH were elevated in some males and females of the 810 mg/kg groups. 6. No abnormalities due to IPD-1151T administration were found in urinalysis, opthalmological examination, electrocardiography, and fecal occult blood examination, or organ weights. 7. Autopsies including histopathological and electron microscopic examinations on the sacrificed dogs revealed no abnormalities. Subserosal hemorrhage in the base of the heart, congestion in the lungs, congestion and vacuolation in the liver and slight cell infiltration around vessels of the brain were found in the female that died.(ABSTRACT TRUNCATED AT 400 WORDS)

[A thirteen-week oral repeated dose toxicity study of suplatast tosilate (IPD-1151T) in rats].

A 13-week oral repeated dose toxicity study of Suplatast tosilate (IPD-1151T), a new anti-allergic agent, as well as a 5-week recovery study were carried out at dose levels of 0 (control), 200, 600, 1800 and 5400 mg/kg/day using male and female rats. The results were as follows: 1. In general conditions, salivation were observed in some rats of both sexes given 1800 mg/kg/day. Both sexes given 5400 mg/kg/day disclosed salivation and soft stool and then died after showing ataxic gait, hyperesthesia and convulsion of legs. 2. Inhibition of body weight gain in both sexes given 5400 mg/kg/day were observed from the early stage of the treatment period. 3. The food consumption was decreased from about 3-week and the water consumption was increased from the initiation of study to about 3-week in both sexes given 5400 mg/kg/day. However, both of them were remarkably decreased prior to death. 4. Fecal examination for occult blood showed an increasing tendency in the incidence of positive findings in both sexes given 1800 mg/kg/day. 5. Hematological examination showed slight decreases in erythrocytic parameters in both sexes given 1800 mg/kg/day. In both sexes given 5400 mg/kg/day hemoconcentration was observed, some animals showing decreases in leucocyte and lymphocyte counts and lymphocyte percentage. 6. Biochemical examination showed increases in total and free cholesterol levels in males given 600 mg/kg/day or more, an increased cholinesterase and decreased levels of triglyceride and cholesterol ester ratio in males given 1800 mg/kg/day. An increase in LDH was observed in both sexes given 5400 mg/kg/day and half of these animals also showed increases in GOT and Urea N. 7. The absolute weights of the pituitary, brain, thymus, heart, lungs and kidneys were increased. However, no histopathological lesion was observed in these organs. As treatment-related histological changes, atrophy in the thymus and spleen, dilation in digestive tracts, neuronal necrosis and necrotic foci in the central nervous system, necrosis of lymphocytes in the lymphoid organs and a decrease in bone marrow cell were observed in both sexes given 5400 mg/kg/day. 8. After a 5-week recovery period, above-mentioned changes had disappeared. 9. From the above results, the non-effective dose level was estimated to be 200 mg/kg/day in males and 600 mg/kg/day in females, and toxic dose level 1800-5400 mg/kg/day in both sexes.

[Reproductive and developmental toxicity study of suplatast tosilate (IPD-1151T) (1)--Fertility study in rats by oral administration].

The effect of suplatast tosilate (IPD-1151T) on reproductive ability and fetal development of the rat was studied. IPD-1151T was administered orally at dose levels of 0 (control), 200, 600, and 1800 mg/kg/day for the premating, mating and early pregnant period. For parent animals, IPD-1151T caused no abnormalities in clinical signs, reproductive ability, or autopsy findings; the females at 1800 mg/kg/day showed a reduction in body weight gain from one week after the start of administration and a decrease in food consumption from the day of the start of administration to day 6 of pregnancy. For fetuses, IPD-1151T produced no abnormalities in external, visceral or skeletal examinations; embryofetal mortality was increased at 1800 mg/kg/day. The results suggest that the non-effective dose level of IPD-1151T is 1800 mg/kg/day for males and 600 mg/kg/day for females in general toxicity, 1800 mg/kg/day for both sexes in reproductive ability, and 600 mg/kg/day for fetuses under the conditions of this study.

[Reproductive and developmental toxicity study of suplatast tosilate (IPD-1151T) (2)--Teratological study in rats by oral administration].

A teratological study of suplatast tosilate (IPD-1151T), a new anti-allergic agent which has a suppressive action on IgE antibody formation, was conducted with pregnant Wistar rats. Dosage levels of IPD-1151T 0, 300, 900 and 2700 mg/kg/day were administered to dams orally by gavage on days 7 through 17 of gestation. Two-thirds of dams per group was caesarean-sectioned on day 20 of gestation and their fetuses were removed for examination of external, visceral and skeletal anomalies. The remaining one-third was allowed to deliver naturally. F1 neonates were examined developmental, functional and behavioral parameters and reproductive abilities. The results were as follows: 1. Toxicities on F0 dams in the 2700 mg/kg/day group were salivation, piloerection, and decreases in body weight and food consumption. Seven animals (19.4%) showed severe toxicity and were dead. Toxicity in the 900 mg/kg/day group was a slight decrease in food consumption. The dosage level of 300 mg/kg/day was non-toxic. 2. Toxicities on F1 fetuses in the 2700 mg/kg/day group were a decrease in body weight and an increase in visceral anomalies (main one was ventricular septal defect that might be related to developmental retardation). No toxicities were seen in the 300 and 900 mg/kg/day. 3. In F1 neonates, suppressions of body weight were observed clearly in the male and female 2700 mg/kg/day and slightly in the male 900 mg/kg/day groups. But no changes in parameters of development, function, behavior or reproductive ability were seen in any dosed groups. It was suggested that no effective dose levels of IPD-1151T were 300 mg/kg/day for F0 dams and F1 neonates, and 900 mg/kg/day for F1 fetuses.

[Reproductive and developmental toxicity study of suplatast tosilate (IPD-1151T) (4)--Perinatal and postnatal study in rats by oral administration].

In order to assess the effect of suplatast tosilate (IPD-1151T) on pregnancy of the rat, and the post natal development to maturity of the F1 generation, daily doses of 0 (control), 200, 600 and 1800 mg/kg/day were administered orally to female Wistar rats from day 17 of pregnancy to day 21 after delivery. All females were allowed to give birth and rear their young to weaning. F0 dams showed no treatment-related changes in general conditions including the state of delivery or nursing, gestation period, birth rate, or autopsy findings. The dams at 1800 mg/kg/day showed a tendency to reduction in body weight gain and a decrease in food consumption. F1 offspring showed no treatment-related changes in clinical signs on the birth day or during the nursing period, external examination on the birth day, general condition, physical or reflex development, open-field test, water multiple T-maze test, autopsy findings, organ weights, skeletal examination, reproductive ability, or histopathological examination on the reproductive organs of the animals of both sexes that failed to produce pregnancy. The offspring at 1800 mg/kg/day showed a reduction or its tendency in body weight gain. There were no treatment-related changes in any of reproductive parameters of F1 dams including external examination of F2 fetuses. The results suggest that the non-effective dose level of IPD-1151T is 600 and 1800 mg/kg/day for F0 dams in general toxicity and reproductive ability, respectively, and 600 mg/kg/day for F1 offspring in post natal development under the conditions of this study.

[Reproductive and developmental toxicity study of suplatast tosilate (IPD-1151T) (3)--Teratological study in rabbits by oral administration].

A teratological study of suplatast tosilate (IPD-1151T), a newly developed anti-allergic agent, was carried out in pregnant NZW rabbits to assess the effects on dams and fetuses. IPD-1151T was administered to dams orally at dose levels of 0, 100, 300, 450 and 700 mg/kg/day from day 6 through day 18 of gestation, and their fetuses were removed for teratological evaluation. The results were as follows: 1. In dams, marked increase in the incidence of abortion, and decrease in body weight gain, food consumption and feces mass were shown in the 700 mg/kg/day group. Slight decrease in body weight gain and food consumption were seen in the 450 mg/kg/day group, but no toxicities were observed in the 300 mg/kg/day or less groups. 2. In fetuses, marked increase in embryo-fetal deaths and decrease in alive fetal body weights and placental weights, but no teratogenicity were shown in the 700 mg/kg/day group. There were no fetal toxicity or teratogenicity in the 450 mg/kg/day or less groups. 3. No effective dose levels were 300 mg/kg/day for maternal general toxicity and 450 mg/kg/day for maternal reproductive toxicity and for fetuses.

[A thirteen-week oral repeated dose toxicity study of suplatast tosilate (IPD-1151T) in dogs].

A 13-week oral repeated dose toxicity study of suplatast tosilate (IPD-1151T), a new anti-allergic agent, as well as a 5-week recovery study were carried out at dose levels of 0 (control), 50, 150, 450 and 1350 mg/kg/day using male and female beagle dogs. The results were as follows: 1. In general conditions, soft feces and diarrhea with specific smell were dose-dependently observed in males and females given 450 mg/kg/day or more. Both sexes given 1350 mg/kg/day, revealed reeling with dropped head, abnormal gait, dysstasia, lying at lateral or prone position, sedation, and tremor, and one male and one female in this group died after showing respiratory depression, collapse and cyanosis. 2. There were no significant or remarkable changes in body weight, food consumption, water consumption, ophthalmology, electrocardiogram, urinalysis, hematology, biochemistry, fecal occult blood test, and absolute and relative organ weights. 3. Pathological examination in dead animals revealed hemorrhagic change in the heart and slight vacuolar changes in hepatocytes. In survived animals, there were no pathological changes attributable to the IPD-1151T. 4. In electron microscopic examination, there were no abnormalities in the liver and kidney attributable to the IPD-1151T. 5. After 5-week recovery period, above-mentioned changes disappeared. 6. From the above results, the non-effective dose level and the toxic dose level were estimated to be 150 mg/kg/day and 1350 mg/kg/day, respectively, and no sex differences were found.

[A fifty-two week oral chronic toxicity study of suplatast tosilate (IPD-1151T) in rats].

A 52-week oral repeated dose toxicity study of suplatast tosilate (IPD-1151T), a new anti-allergic agent, as well as a 5-week recovery study were carried out at dose levels of 0 (control), 50, 300 and 1800 mg/kg/day using male and female rats. The results were as follows: 1. In general conditions, transient salivation after each administration and excretions with peculiar smells were noted in both sexes given 1800 mg/kg/day. Since one male and six female rats given 1800 mg/kg/day showed bradypnea, clonic/tonic convulsions, lying on the belly and/or side, subnormal temperature, abnormal gait, paralysis of extremities, they were sacrificed in moribund. 2. The body weight was lowered from the early stage of administration period in both sexes given 1800 mg/kg/day. 3. There were no remarkable changes in food consumption, urinalysis, fecal examination, hematology and ophthalmology. 4. Biochemical examination revealed a decrease in triglyceride in males given 300 mg/kg/day or more. 5. In pathological examination, the animals sacrificed in moribund showed necrosis and degeneration of neurons and/or sponge-like change of neuropile in nucleus caudatus of the cerebrum, necrosis and partial disappearance of granular cells and Purkinje's cells, and swelling of Bergmann's cells in the cerebellum. In survived animals, the relative organ weight in the liver increased in males given 300 mg/kg/day or more and females given 1800 mg/kg/day, and histopathological examination revealed slight vacuolization, hypertrophy of centrilobular hepatocytes in males given 1800 mg/kg/day. Furthermore, in some females, similar changes of the cerebrum and the cerebellum, as mentioned above, were slightly observed. In electron microscopic examination, slight proliferation of smooth endoplasmic reticulum in hepatocytic cytoplasm was observed in males given 1800 mg/kg/day. The necrobiotic changes, such as condensation of nuclear chromatin, increased electron density of cytoplasm and nuclei, mitochondrial accumulation and vacuolization, in the cells possibly derived from small granular cells in the cerebellum were observed in females given 1800 mg/kg/day. The mitochondrial swelling, decreased and dilated rough endoplasmic reticulum, and increased electron density of cytoplasm and nuclei with formation of cytoplasmic vacuole and membranous degenerated structure in neurons of cerebral temporal lobe cortex were observed in females given 1800 mg/kg/day. 6. In a recovery study, electron microscopic examination revealed a slight degeneration of myelinated nerve fibers in the cerebellum in males given 1800 mg/kg/day. On the contrary, there were no remarkable changes in general condition, body weight and various clinical parameters. It was noted that these changes induced by IPD-1151T seemed to be reversible changes.(ABSTRACT TRUNCATED AT 400 WORDS)