Use of combination therapy with cisplatin and calcitriol in the treatment of Y-79 human retinoblastoma xenograft model.

BACKGROUND: Retinoblastoma is the most common primary malignant intraocular neoplasm of childhood. The poor outcomes of patients with metastatic retinoblastoma have encouraged the search for new therapies. In the current study, the efficacy of combination therapy with calcitriol and cisplatin in athymic mice with subcutaneous Y-79 human retinoblastoma tumours was assessed. METHODS: 60 athymic mice were subcutaneously injected with human Y79 retinoblastoma cells. Animals were randomised into four groups: group 1, 50 microg of cisplatin; group 2, 0.05 microg of calcitriol; group 3, 0.05 microg of calcitriol and 50 microg of cisplatin; group 4, control. The cisplatin was administered once a week, and the calcitriol was given five times a week. RESULTS: There was a significant inhibition of tumour growth in animals treated with the combination therapy of calcitriol and cisplatin as compared with controls and cisplatin alone (p = 0.0001 and p = 0.0041 respectively). In terms of toxicity, serum calcium levels were increased, but there was no mortality and minimal nephrotoxicity in any of the groups. CONCLUSION: The present study shows that cisplatin given in combination with calcitriol may be a viable multidrug therapy option in the treatment of high-risk retinoblastoma.

Countermeasure for space flight effects on immune system: nutritional nucleotides.

Microgravity and its environment have adverse effects on the immune system. Abnormal immune responses observed in microgravity may pose serious consequences, especially for the recent directions of NASA for long-term space missions to Moon, Mars and deep Space exploration. The study of space flight immunology is limited due to relative inaccessibility, difficulty of performing experiments in space, and inadequate provisions in this area in the United States and Russian space programs (Taylor 1993). Microgravity and stress experienced during space flights results in immune system aberration (Taylor 1993). In ground-based mouse models for some of the microgravity effects on the human body, hindlimb unloading (HU) has been reported to cause abnormal cell proliferation and cytokine production (Armstrong et al., 1993, Chapes et al. 1993). In this report, we document that a nutritional nucleotide supplementation as studied in ground-based microgravity analogs, has potential to serve as a countermeasure for the immune dysfunction observed in space travel.

A countermeasure to ameliorate immune dysfunction in in vitro simulated microgravity environment: role of cellularnucleotide nutrition.

Considerable evidence suggests that space travelers are immunosuppressed, presumably by microgravity environmental stresses, putting them at risk for adverse effects, such as opportunistic infections, poor wound healing, and cancer. The purpose of this study was to examine the role and mechanisms of nucleotide (NT) supplementation as a countermeasure to obviate immunosuppression during space travel. The in vitro rotary cell culture system, a bioreactor (BIO), was used to simulate the effect of microgravity and to isolate the neuroendocrine effects inherent to in vitro models. The splenocytes from normal mice were cultured in BIO and control tissue culture (TC) flasks with and without phytohemagglutinin (PHA) for mitogen assays. The culture medium was then supplemented with various concentrations of a nucleosides-nucleotides mixture (NS + NT), inosine, and uridine. Cytokines interleukin (IL)-1beta, IL-2, IL-3, tumor necrosis factor-alpha, and interferon (IFN)-gamma were measured from the supernatant by enzyme-linked immunosorbent assay. In the PHA-stimulated cultures the cellular proliferation in the BIO was significantly decreased as compared with the TC flask cells. BIO-cultured cells in the presence of NS + NT maintained mitogen responses similar to the control TC flask cells. The maintenance of the mitogen response in BIO was observed by the supplementation of uridine and not of inosine. These results are in agreement with our earlier results from unit gravity experiments that showed that pyrimidines are more effective in pleiogenic immunoprotection to hosts. Cytokines IL-1beta, IL-2, and IFN-gamma in the BIO supernatants of cells cultured in the presence of NS + NT had a significantly higher response than the control vessel. Thus, supplemental NT, especially pyrimidines, can confer immune protection and enhance cytokine responses during space travel.

Nutrition beyond nutrition: plausibility of immunotrophic nutrition for space travel.

BACKGROUND AND AIMS: Microgravity has adverse effects on the immune system. We examined the effects of supplemental dietary nucleotides on immune function in ground-based in vivo anti-orthostatic tail-suspended (AOS) mice and in vitro (bioreactor-BIO) analogs of microgravity. METHODS: BALB/c mice were divided into the following three groups: group housed, single isolation, and AOS. Mice were fed either control chow or chow supplemented with RNA or uracil. Immune function was assessed by in vivo popliteal lymph node proliferation (PLN), in vitro PHA-stimulated proliferation of splenocytes and cytokine production. BIO splenocytes were cultured in vitro with/without PHA, a nucleoside-nucleotide mixture (NS/NT) or uridine. The cell proliferation and scanning electron microscopic examination for cells were carried out. RESULTS: PLN response was significantly suppressed in AOS mice (P<0.05) and was restored by RNA and uracil diets. Splenocytes from AOS mice had decreased phytohemagglutinin (PHA)-stimulated proliferation, decreased IL-2 and IFN-gamma cytokine levels (P<0.05). These responses were restored by RNA and uracil diets. In BIO cultures, PHA response was suppressed significantly, and uridine and NS/NT restored the proliferative responses. Scanning electron microscopic analysis of cells cultured in BIO revealed cells with pinched, distorted and eroded membranes. Nucleotide supplementation especially uridine restored normal activated cell surface appearance and ruffling. CONCLUSION: In the microgravity analog environment of AOS and BIO, supplemental nucleotides and especially uracil/uridine have up-regulating and immunoprotective effects with potential as a countermeasure to the observed immune dysfunction in true microgravity.

Temperature dependent eclosion rhythmicity in the high altitude Himalayan strains of Drosophila ananassae.

The circadian pacemaker controlling the eclosion rhythm of the high altitude Himalayan strains of Drosophila ananassae captured at Badrinath (5123 m) required ambient temperature at 21 degrees C for the entrainment and free-running processes. At this temperature, their eclosion rhythms entrained to 12h light, 12h dark (LD 12:12) cycles and free-ran when transferred from constant light (LL) to constant darkness (DD) or upon transfer to constant temperature at 21 degrees C following entrainment to temperature cycles in DD. These strains, however, were arrhythmic at 13 or 17 degrees C under identical experimental conditions. Eclosion medians always occurred in the thermophase of temperature cycles whether they were imposed in LL or DD; or whether the thermophase coincided with the photophase or scotophase of the concurrent LD 12:12 cycles. The temperature dependent rhythmicity in the Himalayan strains of D. ananassae is a rare phenotypic plasticity that might have been acquired through natural selection by accentuating the coupling sensing mechanism of the pacemaker to temperature, while simultaneously suppressing the effects of light on the pacemaker.

Crossed polydactyly.

A common and conspicuous congenital hand anomaly, polydactyly commonly involves only the hand or the foot. Polydactyly involving both hands and feet is rare. We herewith report two cases of Crossed Polydactyly (Type I) and review the literature.

Evaluation of benzalkonium chloride chemoneurolytic proximal gastric vagotomy.

BACKGROUND: Transmucosal chemoneurolytic injection of benzalkonium chloride (BAC) has previously been shown to duplicate operative proximal gastric vagotomy (PGV) in controlling gastric acid secretion. In this study, BAC was evaluated as to efficacious dose, methods of delivery, and systemic toxicities. METHODS: Sham celiotomy, operative PGV controls, transmucosal injections through a gastrotomy, and transserosal injections of BAC (saline controls, 0. 625, 1.25, 2.5, 5.0, 10 mg BAC/kg body wt) were administered to Sprague-Dawley rats. After 3 months the rats underwent Congo red testing (CRT), horseradish peroxidase (HRP) neuronal staining, and necropsy. The color density change of the gastric mucosa from basic to acidic demonstrated by the CRT at the time of necropsy was used to calculate the residual anatomic acid-secreting area. Prior to necropsy, subserosal HRP injections into the anterior and posterior stomach walls assayed vagal neuronal viability via retrograde axonal flow. Results were compared by an ANOVA. RESULTS: The results demonstrated that 1.25-10 mg/kg transmucosal BAC replicated the results of operative PGV; 2.5 mg/kg was found to be the most effective dose. All injection groups including saline controls demonstrated similar diminished vagal retrograde axonal flow by HRP testing consistent with local BAC chemoneurolytic effects. No systemic toxic symptoms were observed after tail vein intravenous BAC 1.25, 2.5, and 5.0 mg/kg. CONCLUSIONS: These efficacy studies have demonstrated BAC's potential utility in the performance of endoscopic transmucosal chemoneurolytic PGV.

Prophylactic effect of dietary glutamine supplementation on interleukin 8 and tumour necrosis factor alpha production in trinitrobenzene sulphonic acid induced colitis.

BACKGROUND: It is well established that glutamine supplemented elemental diets result in less severe intestinal damage in experimental colitis. However, few studies have examined the mode of action of glutamine in reducing intestinal damage. AIMS: To examine the effects of glutamine supplemented elemental diets on the potent inflammatory cytokines interleukin 8 (IL-8) and tumour necrosis factor alpha (TNF-alpha) in trinitrobenzene sulphonic acid (TNBS) induced colitis which presents with both acute and chronic features of ulcerative colitis. METHODS: Sprague-Dawley rats were randomised into three dietary groups and fed 20% casein (controls), or 20% casein supplemented with either 2% glutamine (2% Gln) or 4% glutamine (4% Gln). After two weeks they received intracolonic TNBS to induce colitis. RESULTS: Both Gln groups of rats gained more weight than the control group (p < 0.05) which had progressive weight loss. Colon weight, macroscopic, and microscopic damage scores for the Gln groups were lower than in the control group (p < 0.05). IL-8 and TNF-alpha concentrations in inflamed colonic tissues were lower in the Gln groups than in the control group (p < 0.05), and correlated well with disease severity. Bacterial translocation was lower both in incidence (p < 0.05) and in the number of colony forming units (p < 0.05) for the Gln groups, than in the control group. With respect to all indices studied, the 4% Gln group performed better than did the 2% Gln group. CONCLUSION: Prophylactic glutamine supplementation modulates the inflammatory activities of IL-8 and TNF-alpha in TNBS induced colitis.

Modulation of age-related changes in immune functions of protein-deficient senescence-accelerated mice by dietary nucleoside-nucleotide mixture supplementation.

In the present study we examined the immune-enhancing effect of a nucleoside-nucleotide mixture on the non-specific T-cell immune functions of senescence-accelerated mice (SAM) fed on a low-protein diet. The immune functions studied were in vitro thymic and splenic cell lymphoproliferative responses to phytohaemagglutinin, lipopolysaccharide and concanavalin A and their production of interleukin-2 (IL-2) and interferon-gamma (INF-gamma) in response to mitogen stimulation. SAMP8 mice aged 3 and 6 months were used. In each age group, mice were fed on diets containing either 50 g casein/kg, 50 g casein/kg supplemented with 5 g nucleoside-nucleotide mixture/kg or 200 g casein/kg for 3 weeks. The supplemented 3- and 6-month-old mice had higher (P < 0.05) thymic and splenic cell counts compared with the low-protein group. In both age groups of mice, concanavalin A induced higher (P < 0.05) total thymic and splenic lymphoproliferative responses for the nucleoside-nucleotide mixture-supplemented group compared with the 50 g casein/kg dietary groups. Thymic and splenic production of IL-2 was higher for the 3-month-old mice in both the supplemented and the 200 g casein/kg dietary groups. INF-gamma production in the supplemented 3-month-old group and the 6-month-old 200 g casein/kg dietary group was higher (P < 0.05) compared with the other groups. Overall the supplemented 3-month-old mice exhibited both higher lymphoproliferative responses and production of cytokines compared with the supplemented 6-month-old mice. The results indicate that early nucleoside-nucleotide mixture supplementation may enhance the immune response in protein-deprived SAMP8 mice.

Effect of dietary nucleosides and nucleotides on murine allergic rhinitis.

Although there are studies that report the effects of dietary nucleoside and nucleotide mixtures on the immune response, none are concerned with the role in allergic disease. This study evaluated the effect of dietary nucleic acid mixture (NAM) on mice with a nasal allergy model. One group of mice was supplemented with a 0.5% NAM and the other two groups were fed with a nucleic acid-free diet with 20% casein that served as sensitized and nonsensitized controls. The mice of the NAM group and the sensitized control group were sensitized in two courses by 2 microl of 5% 2,4-toluene diisocyanate (TDI), whereas the nonsensitized control was given 2 microl of ethylacetate instead of TDI. On the 28th day, an allergy was provoked with 4 microl of 2.5% TDI and the allergic responses were observed for 10 minutes. Results showed that the NAM diet group had more severe symptoms of itching, rhinorrhea, snorting, and irritability compared with the controls; also observed were a high incidence of sneezing at 34.7 +/- 4.0 in NAM compared with 19.0 +/- 3.0 (P < 0.001) in sensitized controls and 2.8 +/- 0.7 in nonsensitized controls. From this study, it can be concluded that diets supplemented with nucleic acid mixture contribute to the severity of murine allergic rhinitis.

Nucleoside-nucleotide free diet protects rat colonic mucosa from damage induced by trinitrobenzene sulphonic acid.

BACKGROUND: Growing evidence suggests that intestinal recovery from injury induced by radiation, endotoxin, and protein deficiency is improved by the ingestion of nucleosides and nucleotides. AIM: This study examined the effect of dietary nucleosides and nucleotides supplementation on trinitrobenzene sulphonic acid induced colonic damage in experimental colitis. METHODS: Sprague-Dawley rats were randomised into two groups and fed nucleic acid free 20% casein diet (control) or this diet supplemented with 0.5% nucleoside-nucleotide mixture for four weeks. On the second week, colonic inflammation was induced in rats by intracolonic administration of 0.25 ml of 50% ethanol containing 25 mg of trinitrobenzene sulphonic acid. Additionally, other sets of rats were treated with 0.25 ml of 50% ethanol, 25 mg of trinitrobenzene sulphonic acid in 0.25 ml saline, or 0.25 ml of 0.9% saline. RESULTS: After two weeks, colon weight, macroscopic and microscopic damage scores, were significantly greater (p < 0.05) in the nucleoside-nucleotide supplemented group compared with the non-supplemented control groups. The same variables seen in the trinitrobenzene sulphonic acid-ethanol group fed nucleoside-nucleotide free diet were greater (p < 0.05) than in the rest of the groups fed nucleoside-nucleotide free diet and treated with ethanol, trinitrobenzene sulphonic acid in saline, or saline. Histologically, segmental ulceration and inflammation associated with significantly increased infiltration of polymorphonuclear leucocytes, macrophages, lymphocytes, fibroblasts were observed in the supplemented group compared with the controls. In the nucleoside-nucleotide supplemented group the epithelial damage, mucosal erosion, oedema, and coagulative necrosis of the muscularis propria was more extensive in comparison to the non-supplemented control groups. CONCLUSIONS: This study suggests that dietary nucleosides and nucleotides may aggravate colonic damage and inflammation in chemically induced experimental colitis in rats; and that nucleoside-nucleotide free diet combined with other pharmacological agents may offer a better response.

A nucleoside-nucleotide mixture and its components increase lymphoproliferative and delayed hypersensitivity responses in mice.

We studied the effect of individual components of a nucleoside-nucleotide mixture on T cell-mediated immunity in BALB/c and DBA/2 mice. Mice were fed for 4 wk a nucleotide-free 20% casein diet (control) or this diet supplemented with 3 mol/kg of one of the following; inosine, guanosine monophosphate, uridine, cytidine, thymidine, or a mixture of these. In both strains of mice, popliteal lymph node immunoproliferative response to alloantigeneic (C57BL/6 splenic cells) challenge in mice fed the mixture and guanosine monophosphate was greater (P < 0.05) than in the mice fed the control diet. BALB/c mice fed inosine, uridine, and thymidine also showed greater (P < 0.05) responses compared with control fed mice. In the sheep red blood cells challenge assay, foot pad weight-gain in both strains of mice fed the mixture and the individual components supplemented diets was greater (P < 0.05) than in those fed the control diet. In both strains of mice, interleukin-2 secretion by popliteal lymph node lymphocytes in mice fed the mixture and thymidine was higher (P < 0.05) compared with the rest of the groups except BALB/c mice fed cytidine. Significantly higher than control secretions of interferon-gamma were observed only in BALB/c mice fed inosine, thymidine and the mixture. We conclude that mice fed the nucleoside-nucleotide mixture and the individual components of the mixture had greater responses in the T cell-mediated immune functions studied and that the responses in mice fed the mixture were not different from those in mice fed the individual components.

Evidence for a colonic PAF receptor.

We sought to determine the effect of exogenously administered platelet-activating factor (PAF) on eicosanoid release from the left colon of the rabbit. Using an isolated buffer-perfused rabbit left colon preparation, 1.0- or 5.0-micrograms doses of PAF were infused into the inferior mesenteric artery. Effluents from the inferior mesenteric vein and colonic lumen were collected and the concentrations of the eicosanoids, prostaglandin E, 6-ketoprostaglandin F1 alpha, thromboxane B2, and leukotriene B4 (LTB4), were measured by ELISA. During PAF infusion there was a significant increase of all prostanoids, but not LTB4 into the venous effluent and colonic luminal perfusate when compared to control experiments. Additional studies were performed by pretreating the colons with the PAF antagonists WEB-2170 or alprazolam prior to PAF infusion. Both venous and luminal effluent prostanoid release was effectively blocked by WEB-2170, but not by alprazolam. Colons pretreated with WEB-2170 prior to PAF had markedly diminished tissue injury when compared to colons treated with PAF alone. Inhibition of PAF-stimulated prostanoid release by WEB-2170 suggests that a PAF-sensitive receptor is present in rabbit colonic tissue which may induce eicosanoid-mediated tissue injury.

The role of dietary sources of nucleotides in immune function: a review.

Dietary sources of preformed purines and pyrimidines seem to be important for optimal function of the cellular immune response. It was previously assumed that nucleotides were not needed for normal growth and development, but the results described in this review demonstrate a need for nucleotides in the response to immunological challenges. This effect is likely due to a requirement for preformed pyrimidines for proper development and activation of T cells. The need for sources of preformed nucleotides in defined formulas such as parenteral and enteral formulas and infant formulas is suggested by the studies reviewed below.

The role of nucleotides in adult nutrition.

Dietary nucleotides, found in normal diets, have been recently determined to be required for normal immune defenses. Rejection of cardiac transplants, graft-vs.-host disease, and delayed cutaneous hypersensitivity in animal models are all suppressed by a diet deficient in nucleotides. T lymphocytes seem to require dietary nucleotides for normal maturation and function. Host resistance to bacterial and fungal infections is decreased in mice on nucleotide free diets; addition of RNA or uracil prevents this vulnerability to infection. Dietary RNA is required to restore lost immune function after protein deprivation. Adequate calories and protein alone do not return immune function to normal. Dietary nucleotides can restore lost immune function even during protein starvation and weight loss. Because all parenteral and most enteral nutrient solutions are nucleotide free, clinical studies were undertaken comparing a new nucleotide containing diet (Impact) to a standard high protein enteral feeding. In two separate double blind clinical studies the patients fed the enteral diet containing nucleotides had improved immune function compared with patients receiving a nucleotide free diet. In addition, infectious complications and length of hospital stay were reduced in postoperative cancer patients fed Impact compared with a control group.

Immunohemopoietic effects of dietary nucleotide restriction in mice.

The influence of dietary sources of nucleotides on host in vivo and in vitro immuno-hematologic responses in BALB/c (NCI) mice was studied. Adenosine deaminase (ADA) and purine nucleoside phosphorylase (PNP) were measured in popliteal lymph nodes undergoing proliferative response to syngeneic and allogeneic in vivo stimulation. Supplementation of a nucleotide-free (NF) diet with yeast RNA (NFR) or uracil (NFU) significantly enhanced the host PLN immune response as compared with NF and NF supplemented with adenine (NFA) diets. Levels of ADA and PNP enzymes in the PLNs increased with the alloimmune PLN response of host, and immunosuppression was associated with decreased ADA and PNP activities in lymphocytes following antigenic stimulation. The induction of these enzymes during immune response appears to require dietary sources of certain nucleotides. When bone marrow cells from control chow fed animals were cultured with supernatants (sups) from mitogen activated splenocytes of animals on each dietary group, NF sups significantly decreased (P less than 0.05) the BM proliferative response compared with the response observed with NFR sups, and similar to NFA or NFU sups. When stimulated with purified IL-3, NFR BM cells had higher levels of Thy1.2 or Lyt 1 surface markers as compared with other test groups. In the in vivo splenic colony formation-CFUs assay, spleens from NFR- and NFU-fed animals had a significantly higher number of colonies than spleens from NF- or NFA-fed mice. Thus, NF diet decreases both in vivo lymphoproliferation response to alloantigen and hemopoietic growth factor production, rendering the host splenic environment deficient for stem cell growth. These adverse effects are reversed by RNA supplementation of NF diet. These nutritional studies demonstrate a critical and regulatory role for dietary nucleotides in immunohemopoiesis.

A potential basis for suppressed inflammatory cell function in pediatric cholestatic hosts.

Infective mortality is common in children who have hepatic failure. We have demonstrated that experimental hepatic failure (EHF) profoundly suppresses T cell function in vivo. To determine the basis for immune suppression in EHF we postulated that this phenomenon is attributable to alterations in accessory macrophage (Ma) function, T cell subsets, interleukin-2 (IL-2) production, or serum inhibition. Wistar Furth rats (200 g) were randomized to EHF (n = 23), Sham (n = 23), and normal control (NC) (n = 23) groups. On day 21, splenocytes and sera were harvested and immune assays performed in vitro. Following are the results (mean +/- SEM; Student's t test). Serum bilirubin was elevated in EHF versus Sham and NC groups (P less than .01). EHF splenic macrophages suppressed PHA when added to microcultures at 10(5) concentration (-140 +/- 550 v 12,263 +/- 2,492 [Sham] and 21,413 +/- 1,702 [NC] P less than .01). This effect was not evident when macrophages were added back to microcultures at 10(3) and 10(4) concentrations, suggesting a dose-dependent inhibitory effect. T helper: suppressor ratios did not differ in EHF (1.3 +/- 0.2) compared with Sham (1.4 +/- 0.2) and NC groups (1.2 +/- 0.1). IL-2 production was similar in EHF, Sham, and NC animals (112,141 +/- 5,232 versus 106,691 +/- 1,419 and 120,759 +/- 3,249 counts per minute). T cell inhibitory activity was not demonstrable in EHF sera. These data show that splenic macrophages can inhibit T cell function in vitro. This phenomenon may be paramount in predisposing children with liver disease to infection.