Inhibitory effect of porphyrins on the proliferation of mouse spleen lymphocytes in vitro

The influence of various porphyrins (deuteroporphyrin IX, mesoporphyrin IX, protoporphyrin IX, hematoporphyrin) and two related compounds (hemin, biliverdin) on the spontaneous proliferation of mouse spleen lymphocytes has been estimated in vitro by the 3H-thymidine uptake assay. It has been found that porphyrins (endogenous ligands for the mitochondrial benzodiazepine receptor) produce a concentration-dependent inhibition of 3H-thymidine incorporation into the DNA of these cells. Metalloporphyrin-hemin has been observed to evoke a weak inhibitory effect, in a high concentration (10(-4)M), whereas biliverdin, a porphyrins degradation product, was inactive in the same experimental conditions. Those findings indicate that endogenous porphyrins, presumably acting through the mitochondrial benzodiazepine receptor, could regulate the proliferation of mouse spleen lymphocytes in vitro.     

连翘酯苷对小鼠脾脏淋巴细胞体外增殖与分泌功能的影响

目的分析连翘酯苷（FS）对小鼠脾脏T和B淋巴细胞增殖、分泌NO和TNF-α的影响,初步探讨其免疫调节作用机制。方法无菌操作分离小鼠脾脏,制备脾脏细胞并用含10%胎牛血清的RPMI 1640培养,在培养液中分别加入刺激剂刀豆蛋白（ConA）和脂多糖（LPS）以及不同浓度40、80、160μg/mL的FS共培养不同时间,采用MTT法检测T和B淋巴细胞的吸光度变化,ELISA和Griess法分别检测细胞分泌TNF-α和NO的水平。结果低浓度和中浓度FS对ConA诱导T淋巴细胞24 h和48 h后细胞增殖和存活率明显提高,诱导时间延长至72 h后FS明显抑制细胞转化;低浓度FS对LPS诱导脾脏B淋巴细胞24 h后细胞增殖和生存率显著提高;FS促进小鼠脾脏T和B淋巴细胞分泌NO;FS促进B淋巴细胞分泌TNF-α,中浓度FS促进T淋巴细胞分泌TNF-α而高浓度反而抑制其分泌。此外,FS对环磷酰胺（CY）处理小鼠的脾脏淋巴细胞体外增殖有明显影响,对细胞NO分泌影响不显著。结论结果提示FS可能通过影响小淋巴细胞增殖和细胞因子分泌而调节免疫细胞功能。     

Inhibitory effect of thyrotropin releasing hormone on spontaneous proliferation of mouse spleen lymphocytes in vitro.

The effects of thyrotropin releasing hormone (TRH) and of TRH-like tripeptide pGlu-His-Gly-OH (colon mitosis inhibitor, CMI) on spontaneous proliferation of murine splenocytes were investigated in vitro. The 3H-thymidine incorporation into splenocyte DNA was used as an index of proliferation. It was found that TRH suppressed the proliferation of splenocytes. In contrast, CMI was ineffective by itself but used together with TRH blocked the effect of the latter.     

Effects of pulsed electric fields on mouse spleen lymphocytes in vitro

The effects of pulsed electric fields on cell membranes were investigated. In vitro exposure of mouse splenocytes to a single high-voltage pulse resulted in an increase in membrane permeability that was dependent on both the electric field strength and the pulse duration. Exposure to a 2 μs, 3.0 kV/cm pulse resulted in the induction of a 1.26 V transmembrane potential, and elicited a 50% loss of intracellular K⁺. These results are in agreement with previous studies of the effects of pulsed electric fields on erythrocytes and microorganisms. The effect of pulsed electric fields on the functional integrity of lymphocytes was i vestigated by measuring [³H]thymidine incorporation by cells cultured in the presence and absence of various mitogens following exposure to an electrical pulse. No statistically significant effects on the response of mouse spleen lymphocytes to concanavalin A, phytohemagglutinin or lipopolysaccharide were observed following exposure to 2 μs electric pulses at amplitudes of up to 3.5 kV/cm. Exposure to a single 10 μs pulse of 2.4–3.5 kV/cm produced a statistically significant reduction in the response of lymphocytes to lipopolysaccharide stimulation that was attributed to cell death.     

In vitro screening of seaweed extract on the proliferation of mouse spleen and thymus cell

A total number of 31 types of seaweed were assessed with regard to their effects on the proliferation of mouse spleen and thymus cells in a culture, using an MTT reduction assay. Acetone:dichloromethane (1∶1) extracts of three seaweed plants:Derbesia marina, Sargassum sp., andHisikia fuziformis, exhibited significantly positive effects on the survival of mouse spleen and thymus cellsin vitro. The acetone: dichloromethane (1∶1) extracts ofSargassum sp., in particular, much more potent effects on thymus cell activation than did any of the other types of seaweed. However, the methanol extracts ofSargassum ringgoldianium andChondrus crispus exerted a stimulatory influence only on the proliferation of mouse spleen cells, whereas the methanol extracts ofGrateloupia lanceolata exhibited significant cell proliferation properties in both spleen and thymus cells.     

Effect of lymphocytes on the differentiation of in vitro transformed L-line mouse fibroblasts

Possible participation of immune system cells in the differentiation of non-lymphoid tissues has been examined. Lymphocytes were tested on transformed fibroblasts of L mouse strain. The presence of lymphocytes in L cell culture enhanced their differentiation, assessed by morphological, proliferative and biophysical criteria. The induction of L-cell differentiation took place during contact and short-distant interaction both with mouse and human lymphocytes. The phenomenon had a stable character, as it was revealed in L cells separated from lymphocytes during passaging. The results obtained indicate real and potential abilities of the immune system to affect differentiation of proliferating non-lymphoid tissues.     

Effect of beta-endorphin and myelopeptides on the cAMP level and proliferation of lymphocytes in vitro

Beta-endorphin (10(-11)-10(-9) M) has been shown to induce naloxone-independent depression of the proliferative activity of human peripheral lymphocytes (HL), stimulated by pokeweed mitogen without affecting PHA-stimulated HL proliferation. Beta-endorphin (10(-10)-10(-7) M) also caused changes in HL cAMP level, that were blocked by naloxone. Marked individual sensitivity to beta-endorphin effects has been noted. It has been also shown that a bone marrow preparation, stimulating antibody production (myelopeptides), causes naloxone-independent depression in the proliferative activity of HL, stimulated by PHA and pokeweed mitogen, as well as naloxone-blocked decrease in cAMP HL level. It has been concluded that beta-endorphin interacts with several types of opiate lymphocyte receptors and that opioids, contained in myelopeptides, are involved in the realization of myelopeptide effect on lymphocytes.     

Heterogeneous uptake of 9-beta-D-arabinofuranosyladenine by mouse spleen lymphocytes

Spleens of mice treated with 9-beta-D-[2,8-3H]arabinofuranosyladenine (araA), 0.5-30.0 mg/kg showed rapid dose-dependent accumulation of 3H, which peaked at 2.5 mg/kg. Lymphocytes from spleens showed linear uptake when incubated with 0.68, 1.36, and 5.03 microM araA over 120 s. With 1.0 mM araA, uptake was reduced in rate but was not saturated. Tritiated metabolites identified from these lymphocytes were araA which predominated after brief incubations, its deaminated form (araH), and some phosphorylated product in small amount. Inhibition of deaminase increased intracellular araA. Although potent inhibitors of nucleoside transport inhibited araA uptake marginally, adenosine competed with araA for about 50% of the uptake capacity. The data suggest that the uptake of araA by mouse lymphocytes, which is not simple diffusion, occurs by a mechanism distinct from typical nucleoside transport.     

Response of spleen lymphocytes shortly after thymectomy in adult mouse.

The blastogenesis response to the phytomitogens, PHA-P, Con A and PWM was used to assess the effect of adult thymectomy on the spleen lymphocytes of C57B1 mice. The mitogenic response to the phytomitogens was determined by 3H-thymidine uptake. The changes produced in theta-antigen bearing spleen lymphocytes were also evaluated making use of theta antibodies from AKR/S mice previously injected with splenic and thymic lymphocytes from CBA/J mice. The present results show that the response to mitogens PHA-P and Con A is reduced early after thymectomy while the response to PWM was only slightly reduced. There was not any correlation between the disminished response to mitogens and the changes observed in theta bearing spleen lymphocytes.     

Dopamine increases cyclic AMP concentration in the rat spleen lymphocytes in vitro

The effect of dopamine on the cyclic AMP concentration in the rat spleen lymphocytes has been investigated $\text{in}\text{vitro}$. It has been shown that dopamine in concentration above 10^?6M induces a significant increase of cyclic AMP level. The maximal stimulatory effect was observed after 10 minutes of the lymphocytes incubation with dopamine. These data suggest that the dopamine receptor in lymphocyte belongs to D-1 category.     

Effect of thymalin on the cyclic nucleotide system in the mouse spleen

The effect of the thymus preparation thymalin (1000 micrograms/mouse, i.p.) on the system of cyclic nucleotides in the spleen has been studied. The preparation produced two-phasic changes in the content of cAMP and cGMP, as well as in the activity of cAMP phosphodiesterase. The changes in the spleen cAMP content are consequent on those in the activity of phosphodiesterase.     

Regulation of glutathione levels in mouse spleen lymphocytes by transport of cysteine

Cysteine and cystine transport activities of resting and activated mouse spleen lymphocytes were characterized in order to examine the contributions of cysteine and cystine to intracellular glutathione contents. Following stimulation with lipopolysaccharide, the lymphocytes markedly increased their capacity to transport cysteine. The uptake of cysteine was mediated mainly by the ASC system (Na+-dependent neutral amino acid transport system especially reactive with alanine, serine, and cysteine). On the other hand, both the resting and the activated lymphocytes had extremely low cystine transport activities. Because of the instability of cysteine, the culture media usually contained cystine but not cysteine. Therefore, both the resting and the activated lymphocytes rapidly decreased their glutathione contents owing to their poor capacities to take up cystine. The effects of freshly added cysteine on the cellular glutathione contents were examined in the presence of bathocuproinedisulfonate, a nontoxic copper-specific chelator that inhibits autoxidation of cysteine. Cysteine added at 25-400 microM only partially prevented the rapid decrease of the glutathione contents in fresh resting lymphocytes. In the lipopolysaccharide-activated cells, however, cysteine enhanced the cellular glutathione contents in a dose-dependent manner. These results indicate that the enhanced activity of the ASC system increases the level of intracellular glutathione in the presence of cysteine.