Lymphocyte Proliferation In Rats Research Articles

Boron Affects Immune Function Through Modulation of Splenic T Lymphocyte Subsets, Cytokine Secretion, and Lymphocyte Proliferation and Apoptosis in Rats.

This study demonstrated the mechanisms of boron effects in a rat model and provided a scientific basis for the rational of boron use. These findings were achieved by investigating the effects of boron (10, 20, 40, 80, 160, 320, and 640mg/L in drinking water or 1.5, 3, 6, 12, 24, 48, and 96mg/kg BW) on rat serum immunoglobulins (IgGs), splenic cytokines, lymphocyte subsets, as well as on lymphocyte proliferation and apoptosis. Addition of 20 (3) and 40 (6)mg/L (mg/kg BW) of boron to drinking water significantly increased rat serum IgG concentrations, splenic IFN-γ and IL-4 expression as well asthe number of splenic CD3+, CD4+ and proliferating cell nuclear antigen (PCNA)+ cells. Supplementation of drinking water with 40mg/L (6mg/kg BW) boron also markedly increased splenic IL-2 expression and the CD4+/CD8+ cell ratio and reduced splenic CD8+ cell number. Supplementation with 80mg/L (12mg/kg BW) boron significantly increased CD3+ and PCNA+ cell numbers (P<0.05) and decreased the IL-10 expression in the spleen. Addition of 320 (48) and 640 (96) mg/L (mg/kg BW) boron markedly reduced the serum IgG concentrations; splenic IL-2 and IL-10 expression; the number of CD3+, CD4+ and PCNA+ cells; and increasedthe number of splenic CD8+ and caspase-3+ cells and promoted caspase-3 expression in CD3+ cells. In conclusion, these findings suggest that the supplementation of rat drinking water with 20(3) and 40(6) mg/L (mg/kg BW) boron can markedly enhance humoral and cellular immune functions, while boron concentrations above 320mg/L (48mg/kg BW) can have an inhibitory effect or even toxicity on immune functions. These results exhibit a U-shaped response characteristic of low and high doses of boron supplementation on immune function and imply that proper boron supplementation in food for humans and animals could be used as an immunity regulator.

Diverse age-related effects of Bacopa monnieri and donepezil in vitro on cytokine production, antioxidant enzyme activities, and intracellular targets in splenocytes of F344 male rats

Aged people are more prone to developing neurodegenerative and infectious diseases, autoimmune disorders, and cancer due to impairment of neuroendocrine-immune functions. Neuronal degeneration and immunosuppression aided by increased generation of reactive oxygen species combined with loss of antioxidant enzyme activities promote the aging process. Bacopa monnieri (brahmi), an Ayurvedic herb, and donepezil, a reversible acetylcholinesterase inhibitor, have been used to reverse cognitive dysfunctions in several neurodegenerative diseases. The aim of this study was to investigate the effects of in vitro incubation of lymphocytes from spleens of young (3-month-old), early middle-aged (8- to 9-month-old), and old (18-month-old) F344 rats with brahmi (0.001%, 0.01%, 0.05%, 0.1%, and 1%) and donepezil (5, 10, 25, 50, and 100μg/ml) on Concanavalin (Con A)-induced proliferation of T lymphocytes and cytokine production, and the activities of antioxidant enzymes [superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glutathione-S-transferase (GST)]. In addition, the effects of these compounds on the expression of intracellular signaling pathway markers (ERK, p-ERK, CREB, p-CREB, Akt and p-Akt), nitric oxide (NO) production, and the extent of lipid peroxidation were measured in the splenocytes. Age-related decline in Con A-induced proliferation of T lymphocytes was not reversed by treatment with brahmi and donepezil but donepezil alone further reduced the lymphocyte proliferation in young rats. Lower doses of brahmi treatment reversed the age-related decrease in Con A-induced IL-2 and IFN-γ production by the splenocytes while their production by splenocytes was suppressed by treatment with donepezil in the young and early middle-aged rats. An age-associated decline in the activities of SOD, CAT, GPx, and GST was evident in the lymphocytes of spleen. Brahmi enhanced CAT activity of lymphocytes in all the age groups while donepezil increased SOD activity in old rats. Both brahmi and donepezil increased GPx and GST activities in a dose-dependent manner in the lymphocytes of all age groups. There was an age-related decline in NO production and increase in the extent of lipid peroxidation in the splenocytes. Brahmi and donepezil increased NO production in the lymphocytes of early middle-aged and old rats. Brahmi reversed the age-related increase in lipid peroxidation in the splenocytes of both early-middle-aged and old rats while donepezil suppressed lipid peroxidation only in the splenocytes of old rats. The expressions of p-ERK1/2 and p-CREB in the splenocytes were elevated following treatment with brahmi and donepezil in the early middle-aged and old rats while age-related decline in p-Akt expression was reversed by treatment of lymphocytes with brahmi alone in early-middle-aged and old rats. Taken together, these results suggest that both brahmi and donepezil exert distinct age-related effects on the cell-mediated immune responses through selective modulation of antioxidant enzyme activities and intracellular targets that may influence the therapeutic efficacy of these drugs in neurodegenerative diseases.

Red blood cell on lymphocyte proliferation in rats with chronic unpredictable mild stress

To study the changes of rat red blood cells on T, B lymphocytes proliferation in chronic unpredictable mild stress (CUMS). Thirty Wistar rats were averagely divided into control group and the CUMS group. The relevant behavioral performance and red blood cells on lymphocytes proliferation Changes by MTT colorimetric were observed. After fourteen days, the growth of weight and organic coefficient of brain decreased in model group (P < 0.05), with a significantly reduced consumption and preference of sucrose solutions, and increased pure water consumption as compared with control group (P < 0.05). Plasma corticosterone levels peaked at seven days but on a declining trend after fourteen days. The rate of red blood cells on T, B lymphocyte proliferation in depression group was lower than non-stress in 7, 14, 21 days (P < 0.05). The repeated chronic mild stress stimulation can induce long-time changes in acts and activities and decrease the rate of red blood cells on T, B lymphocytes proliferation, which can provide a new experimental method to observe the red blood cell immune function under the stress.

Flaxseed, olive and fish oil influence plasmatic lipids, lymphocyte migration and morphometry of the intestinal of Wistar rats

Evaluate the effect of flaxseed, olive and fish oil on the lipid profile, preservation of villosities and lymphocyte migration in the intestinal mucosa of Wistar rats. Thirty Wistar male rats were divided into four groups, which received the AIN-93M diet, with changes only to their lipid source: flaxseed, olive, fish, and soy oil (control group). The serum was separated for the biochemical parameter analysis. A histological evaluation was performed in the ileal portion. The group which was fed fish oil presented lower values when compared to the other treatments for Total Cholesterol, High-density Lipoprotein Cholesterol and Triacylglycerol (p<0.05). The animals treated with fish and olive oils presented better intestinal villosities preservation. Less deposition of lymphocytes was observed in the flaxseed group (p<0.001). This study demonstrated that flaxseed, olive and fish oils present different responses than soy oil for the intestinal mucosa preservation and lymphocyte proliferation in Wistar rats.

Effects of angiotensin-II receptor blockers on experimental autoimmune myocarditis

Background The effects of angiotensin-II type 1 receptor blockers (ARBs) on the treatment of hypertension, heart failure, and other cardiovascular diseases have been confirmed extensively. However, recent studies have emphasized the nonhemodynamic effects of these drugs. The purpose of this study was to investigate the effects of ARBs on the development of experimental autoimmune myocarditis (EAM), and to clarify the mechanisms involved. Methods EAM model was induced in Lewis rats by injection of porcine cardiac myosin subcutaneously. We administered valsartan (a new ARB) to rats with EAM and measured blood pressure regularly. Echocardiography was performed to examine the cardiac function and heart structure of the rats. The severity of myocarditis was detected by histopathological evaluation. We evaluated antigen-specific T-cell proliferation responses to cardiac myosin by the lymphocyte proliferation assay and measured serum levels of Th1 and Th2 cytokines by enzyme-linked immunosorbent assay. Results There was no significant difference in the blood pressure (BP) level between the groups and cardiac function of valsartan-treated rats was significantly improved compared with untreated rats. Valsartan markedly reduced the severity of myocardial lesions and suppressed lymphocyte proliferation in rats immunized with myosin. After drug administration, Th1 cytokines (IFN-γ and IL-2) were significantly down-regulated, while Th2 cytokines (IL-4 and IL-10) were detected to undergo up-regulation. Conclusions The results suggest that valsartan can ameliorate EAM independent of BP-lowering effects. Some of the beneficial effects of ARBs may be due to their immunomodulatory reactions in the modification of helper T-cell balance.

Effects of acute exhaustive physical exercise upon glutamine metabolism of lymphocytes from trained rats

Transitory immunosupression is reported after intense exercise, especially after an increase in training overload and in overtraining. The influence of intense exercise on plasma hormones and glutamine concentration may contribute to this effect. However, the effect of such exercise-induced changes upon lymphocyte and glutamine metabolism is not known. We compared glutamine metabolism in lymphocytes in sedentary (SED) and trained rats. Rats from the moderate group (MOD) swam for 6 weeks, 1 h/day, in water at 32 ± 1 °C, with a load of 5.5% body weight attached to the tail. Animals from the exhaustive group (EXT) trained like MOD, with training increasing to 3 times 1 h a day during the last week, with 150 min rest between each bout. Animals were killed immediately after the last training bout. We observed reduced concentrations of plasma glucose ( p < 0.05), glutamine ( p < 0.05), glutamate ( p < 0.05) in EXT compared to SED. In MOD, decreases in glutamine ( p < 0.05) were observed. Analyzing lymphocyte metabolism, we observed an increase in lactate production and glutamine consumption ( p < 0.05) in MOD ( p < 0.05) compared to SED and a decrease in glutamine consumption ( p < 0.05) and aspartate production in EXT. An increase in the proliferative response of lymphocytes in MOD and EXT was also observed when stimulated by ConA and LPS similarly to SED. Acute exercise promoted decreased glutamine plasma concentration and changes in glutamine metabolism that did not impair lymphocyte proliferation in exhaustive trained rats.

Buprenorphine produces naltrexone reversible alterations of immune status

Substantial evidence demonstrates that administration of high efficacy μ opioid agonists such as morphine modulate the immune response in a dose-dependent and pharmacologically specific manner, indicating functional interactions between the opioid and immune systems. In contrast to the well-characterized immunomodulatory effects of high efficacy μ opioids, little is known about how these effects generalize to other clinically employed opioids and agonists of varying degrees of μ opioid receptor stimulation. Buprenorphine is a μ opioid agonist of intermediate efficacy that is used clinically for pain management and has recently been approved for the treatment of opioid dependence. Recent evidence indicates pharmacological and mechanistic differences between buprenorphine and morphine. Therefore, the aim of the present study was to investigate whether buprenorphine also possesses immunomodulatory properties. The results demonstrate that buprenorphine dose-dependently suppresses splenic natural killer cell activity, lymphocyte proliferation and IFN-γ production in rats in a naltrexone reversible manner, demonstrating pharmacological specificity of buprenorphine-induced immune alterations.

Effects of tramadol on T lymphocyte proliferation and natural killer cell activity in rats with sciatic constriction injury. (National Cheng Kung University, College of Medicine, Tainan, Taiwan) Pain. 2001;92:63–69.

This study investigated the effects of acute and chronic tramadol treatment on T lymphocyte function and natural killer (NK) cell activity in rats receiving chronic constriction injury (CCI) of the sciatic nerve. T lymphocyte function was evaluated based on concanavalin‐A (ConA)‐induced and phytohemagglutinin (PHA)‐induced splenocyte proliferation. NK cell activity was measured by lactic acid dehydrogenase release assay. The effects of tramadol on thermal hyperalgesia were also assessed by measuring paw withdrawal latency (PWL) in the rats. PWL was dose‐dependently reversed by tramadol after acute treatment (single subcutaneous injection) with 10, 20, and 30 mg/kg, respectively. There was no significant change among acute treatments groups in NK cell activity, whereas splenocyte proliferation induced by ConA and PHA was significantly suppressed starting from a dose of 20 mg/kg. The reversal of the thermal hyperalgesia persisted throughout a period of chronic tramadol treatment of 40 and 80 mg/kg per day, respectively, with continuous subcutaneous infusion for 7 days at a uniform rate via osmotic minipumps. No modulation of NK cell activity was found in either dose group. However, the activity of splenocyte proliferation was decreased in the 80 mg/kg per day group when compared with the saline and 40 mg/kg per day groups. Suggest that tramadol treatment has an immunological profile different from pure μ‐opioid agonists like morphine, which is known to suppress both NK cell activity and T lymphocyte proliferation at a subanalgesic dose in CCI rats. Conclude that tramadol treatment may be a better choice than morphine for treatment of chronic neuropathic pain, particularly in patients with compromised immunity.

Effects of tramadol on T lymphocyte proliferation and natural killer cell activity in rats with sciatic constriction injury

We investigated the effects of acute and chronic tramadol treatment on T lymphocyte function and natural killer (NK) cell activity in rats receiving chronic constriction injury (CCI) of the sciatic nerve. T lymphocyte function was evaluated based on concanavalin-A (ConA)- and phytohemagglutinin (PHA)-induced splenocyte proliferation. NK cell activity was measured by lactic acid dehydrogenase release assay. The effects of tramadol on thermal hyperalgesia were also assessed by measuring paw withdrawal latency (PWL) in the rats. PWL was dose-dependently reversed by tramadol after acute treatment (single subcutaneous injection) with 10, 20, and 30 mg/kg, respectively. There was no significant change among acute treatment groups in NK cell activity, whereas splenocyte proliferation induced by ConA and PHA was significantly suppressed starting from a dose of 20 mg/kg. The reversal of the thermal hyperalgesia persisted throughout a period of chronic tramadol treatment of 40 and 80 mg/kg per day, respectively, with continuous subcutaneous infusion for 7 days at a uniform rate via osmotic minipumps. No modulation of NK cell activity was found in either dose group. However, the activity of splenocyte proliferation was decreased in the 80 mg/kg per day group when compared with the saline and 40 mg/kg per day groups. These data suggest that tramadol treatment has an immunological profile different from pure μ-opioid agonists like morphine, which is known to suppress both NK cell activity and T lymphocyte proliferation at a subanalgesic dose in CCI rats. Considering analgesic and immunosuppressive effects, tramadol treatment may be a better choice than morphine for treatment of chronic neuropathic pain, particularly in patients with compromised immunity.

Mechanisms of cocaine-induced decreases in immune cell function

Cocaine has previously been shown to decrease mitogen-induced T lymphocyte proliferation in rats following intravenous administration. However, in this report, it is demonstrated that central administration of cocaine (1–50 μg) had no effect on lymphocyte proliferation responses. Similarly, the quaternary derivative, cocaine methiodide, also suppressed lymphocyte proliferation only when administered peripherally (6.5 mg/kg), and not centrally (1–20 μg). These results suggest that the effects of cocaine were mediated through a peripheral mechanism. Since significant elevations in plasma corticosterone were observed with all routes of administration of cocaine, the effects of cocaine did not appear to be due entirely to activation of the HPA axis. Instead, the peripheral administration of the local anesthetic, lidocaine (5 mg/kg) or the monoamine reuptake inhibitor, RTI-55 (2–5 mg/kg), produced significant suppressive effects on proliferation, suggesting that both of these peripheral activities of cocaine may be involved in the alteration of lymphocyte responses.

Parenteral supplementation with a fish-oil emulsion prolongs survival and improves rat lymphocyte function during sepsis

Nutritional intervention with ω-3 fatty acids during trauma and infection has been shown to improve the clinical outcome of patients and the survival rate in laboratory animals. This study evaluated the effects of parenteral administration of lipid emulsions containing fish oil (FO) or soybean oil (SBO) on survival and T-lymphocyte response during sepsis. Male Sprague-Dawley rats (250–275 g) were prepared for parenteral feeding 4 d before inducing sepsis by cecal ligation and puncture (CLP). Standard resuscitation was provided with normal saline. Thirty minutes after completing CLP, sham control or CLP rats were infused continuously with saline or a parenteral diet containing SBO or a 1:1 FO:SBO emulsion. The survival rate was significantly improved in rats receiving the FO-supplemented diet, with 50% alive by 120 h in comparison with the saline-infused, chow-fed rats (0% alive by 120 h) or the SBO-fed rats (12% alive at 120 h). The T-lymphocyte response was evaluated at 24 h after CLP. Sepsis led to a decline in lymphocyte proliferation in rats infused with saline or the SBO emulsion, which was associated with a greater release of splenocyte interleukin-10, transforming growth factor-β and prostaglandin E 2. Administering the 1:1 FO:SBO parenteral diet during sepsis improved the survival rate and prevented the sepsis-induced suppression of lymphocyte proliferation and interleukin-2 release. The FO effect on lymphocyte function was associated with decreased splenocyte release of transforming growth factor-β and prostaglandin E 2.

Influence of Unsaponifiable Matter from ghee on lymphocyte proliferation and erythrocyte fragility in rats

Cholesterol Oxidation Products (COPs) are implicated in diseases such as atherosclerosis and cancer and are known to affect vital functions such as cell growth and proliferation, and membrane structure and function. Ghee, commonly used in Indian diet contains a considerable amount of COPs and hence has been implicated as a causative factor for cardiovascular diseases. Little or no information is available on the effect of ghee or its constituents on membrane structure and function. In this study, Unsaponifiable Matter(USM) from ghee was dissolved in groundnut oil and force fed to male wistar rats for a period of 8 days. The control rats were fed Groundnut oil (GNO) while the experimental rats were fed with USM derived from either fresh ghee or oxidized ghee. The effect of feeding USM from ghee on lymphocyte proliferation, erythrocyte fragility, erythrocyte lipid composition and platelet aggregation was studied. Spleen weight and lymphocyte proliferation decreased by 30–41 % in the experimental rats. A decrease in the erythrocyte fragility was observed in the ghee fed rats. However, there was no change in erythrocyte ghost membrane lipid composition, platelet aggregation, or in malondialdehyde content, a measure of the “release reaction” in platelets. It is inferred from these studies that the USM from oxidized ghee is immunosupressive, and has no effect on platelet aggregation.

Effects of decreased plasma glutamine concentrations on peripheral lymphocyte proliferation in rats.

The relationship between exercise-induced lowering of plasma glutamine concentrations and proliferation of peripheral lymphocytes was investigated in male Wistar rats. The T-lymphocyte proliferative responses to the mitogen, concanavalin A, were determined by incorporation of radiolabelled thymidine into the DNA in vitro. The rats ran 2 h x day(-1), 6 days x week(-1) for 4 weeks. Analysis immediately after the final period of exercise showed T-lymphocyte proliferation to be significantly depressed, together with a marked decrease in plasma glutamine concentrations. There were also significant increases in serum corticosterone concentrations immediately after exercise. However, following 24-h recovery, this exercise-induced immunosuppression was not statistically significant when compared with the age-matched control group. In the second experiment, in order to clarify the importance of glutamine for immunological function in vivo, methionine sulfoximine, an effective inhibitor of glutamine synthetase was injected intraperitoneally (12.5 mg x kg body mass[-1]). Plasma glutamine concentrations were decreased 4 h after the injection, compared with the placebo control group, and this resulted in a significant decrease in the rate of T-lymphocyte proliferation. This treatment had no effects on serum corticosterone concentrations. These results would suggest that the chronic exercise-induced reduction in proliferation of peripheral T-lymphocytes is a transient reversible phenomenon, which returns to normal levels within 24 h of the final training period. It is also conceivable that this exercise-induced immunosuppression is associated with a decrease in circulating glutamine concentrations.

Gonadectomy impairs lymphocyte proliferation and macrophage function in male and female rats. Correlation with key enzyme activities of glucose and glutamine metabolism

Gonadectomy impairs lymphocyte proliferation and macrophage function in male and female rats. Correlation with key enzyme activities of glucose and glutamine metabolism

Gonadectomy impairs lymphocyte proliferation and macrophage function in male and female rats. Correlation with key enzyme activities of glucose and glutamine metabolism.

The effect of gonadectomy on lymphocyte proliferation and macrophage function (hydrogen peroxide production and phagocytosis capacity) of male and female rats was examined and the results correlated with the activities of hexokinase, glucose-6-phosphate dehydrogenase, citrate synthase and phosphate-dependent glutaminase. Also, the reversion of the changes by the treatment with oestrogen or progesterone or a combination of both was addressed. Taken as a whole, ovariectomy reduced hydrogen peroxide production and phagocytosis capacity by macrophages and also lymphocyte proliferation. Castration of male rats reduced the proliferation of lymphocytes and raised macrophage phagocytosis capacity. The effects on macrophage function were correlated with changes in glucose metabolism, particularly, in the activity of glucose-6-phosphate dehydrogenase.

The Effect of Cold Stress on Lymphocyte Proliferation in Fetal Ethanol-Exposed Rats

The Effect of Cold Stress on Lymphocyte Proliferation in Fetal Ethanol-Exposed Rats

In Vitro Effects of Melatonin on Mitogen-Induced Lymphocyte Proliferation and Cytokine Expression in Young and Old Rats

Melatonin (MLT) treatment in vivo has been shown to have immunomodulatory and anti-immunosenescent effects in the mouse model. In the present report, the in vitro effect of MLT on mitogen-induced lymphocyte proliferation and cytokine expression was evaluated in a rat model. Splenic lymphocytes were isolated from young (6 months) and old (24 months) F344 rats and were incubated with MLT in the presence or absence of mitogens. The proliferative response to concanavalin A (ConA) or PMA plus ionomycin was measured in splenocytes or T cells isolated from young and old rats. In addition, the induction of interleukin-2 (IL-2) and interferon-gamma (IFN-γ) production was measured in MLT-treated and untreated lymphocytes isolated from young and old rats. The ConA-induced lymphocyte proliferation and IL-2 expression were significantly lower and induction of IFN-γ production was significantly higher in splenocytes and purified T cells isolated from old rats compared to splenocytes and T cells isolated from young rats. Treatment of lymphocytes with MLT did not significantly alter ConA-induced lymphocyte proliferation or IL-2 or IFN-γ expression in lymphocytes isolated from either young or old rats. On the basis of these data, we conclude that in vitro MLT treatment had no immunomodulatory effect on lymphocytes from rats.

Potential role of the autonomic nervous system in the immunosuppressive effects of acute morphine administration

These studies investigated the role of the autonomic nervous system in mediating the immunosuppressive effect of morphine on blood lymphocyte proliferation in rats. To determine the contribution of the autonomic nervous system, rats were pretreated with the ganglionic blocker chlorisondamine (5 mg/kg) prior to morphine (7 mg/kg) administration. Ganglionic blockade with chlorisondamine completely antagonized the inhibitory actions of morphine, suggesting that intact ganglionic transmission was required for the inhibition to occur. Blockade of postganglionic parasympathetic neurotransmission with atropine methylbromide (1 mg/kg) or blockade of sympathetic neurotransmission with the α-adrenoceptor antagonist phentolamine (1 mg/kg) did not attenuate the suppressive effect of morphine. Blockade of β-adrenoceptors with propranolol (2.5 mg/kg) resulted in partial antagonism, but this action was not shared by the peripherally acting β-adrenoceptor antagonist nadolol (6 mg/kg). These results suggest that the inhibitory effect of morphine on blood lymphocyte proliferation may be mediated through activation of the autonomic nervous system; however, individual blockade of either the parasympathetic or sympathetic division of the autonomic nervous system was not sufficient to antagonize this immunosuppressive effect.

Effects of caloric restriction on age-related oxidative modifications of macromolecules and lymphocyte proliferation in rats

Decreased immune function associated with aging has been demonstrated in both humans and animals. We hypothesize that reactive oxygen species (ROS)-mediated damage to biological macromolecules may contribute to compromised immune response during aging. In this study, we compared the levels of lipid peroxidation and oxidatively modified proteins in plasma and splenocytes, and the mitogen-induced T lymphocyte proliferation in ad lib-fed (AL) and caloric restricted (CR) Fischer 344 × BNF 1 male rats at the ages of 5, 18, and 31 months. The results show that AL rats exhibit an age-related decrease in proliferative response of splenic lymphocytes to phytohemagglutinin (PHA) and concanavalin A (Con A). This functional decline in T-lymphocytes during aging is inversely correlated to the levels of both lipid peroxidation and protein carbonyl in the plasma and splenic lymphocytes. Caloric restriction, however, can partially reverse the age-dependent decrease in T lymphocyte proliferation and significantly reduce lipid peroxidation and protein carbonyl contents in plasma and splenocytes. The above observations support the hypothesis that the age-associated declines in immune function are related to the oxidative modification of biological macromolecules, which in turn may lead to enzyme inactivation, membrane disruption, and cell senescence. One of the mechanisms by which caloric restriction reverses declined immune function in aged rats is hypothesized to be through reduction in ROS production and thereby protection of cellular macromolecules against oxidative damage.

Optimizing whole blood lymphocyte proliferation in the rat

Use of whole blood for lymphocyte proliferation experiments have advantages over the usual isolated lymphocyte procedure. We have systematically optimized methodology for whole blood lymphocyte proliferation in male Sprague-Dawley rats. Both phytohemagglutinin (PHA) and concanavalin A (ConA) stimulate lymphocyte blastogenesis in whole blood. The response of rat lymphocytes to ConA at supra-optimum conditions doubled that of PHA in contrast to what is observed in human lymphocytes. Proliferation rates relate non-linearly to concentrations of both mitogens with profiles showing lag, log and decline phases with increasing concentrations. The effect of incubation period followed a similar pattern with an optimum growth rate at 72 h. Whole blood from adrenalectomized rats gave higher proliferation rates than blood from intact normal rats. Advantages of the technique include: use of as little as 8 μl of blood for each determination; rats need not be killed; the technique is less cumbersome, less time consuming, and cheaper to carry out than the traditional isolated lymphocyte method.