Computational studies on conditions of the emergence of autopoietic protocells

It has been pointed out that the acquisition of self-maintaining protocells was one of the most important evolutionary steps in the earliest stage of life. However, there remains little evidence to show what the components of the protocells were and how they were acquired. A theoretical study to investigate the possible process of the emergence of protocells is therefore required. In this paper, we present a computational model that demonstrates the emergence and evolution of self-maintaining and self-reproducing structures to analyze conditions under which precellular autocatalytic molecules could evolve into self-reproducing protocells. We focused on the supply rates of molecular resources as environmental parameters and explored the pathways of evolution from molecular replication to cellular reproduction under various conditions. The results showed that the spontaneous organization of protocells from a random initial state takes place in a parameter region where the metabolism becomes difficult because of an insufficient supply of resources, but once cell structures are organized, they can survive in a wider range of environments. We investigated the evolution in temporally or spatially changing environments and found that the protocells can take over the precellular metabolic system after the transition. These results suggest a possible scenario for the evolution from precellular to cellular reproduction.     

IL-7 exacerbates chronic colitis with expansion of memory IL-7Rhigh CD4+ mucosal T cells in mice

We have previously demonstrated that mucosal CD4(+) T cells expressing high levels of IL-7 receptor (IL-7R(high)) are pathogenic cells responsible for chronic colitis. Here we investigate whether IL-7 is directly involved in the expansion of IL-7R(high) memory CD4(+) mucosal T cells and the exacerbation of colitis. We first showed that CD4(+) lamina propria lymphocytes (LPLs) from wild-type, T cell receptor-alpha-deficient (TCR-alpha(-/-)), and recombinase-activating gene (RAG)-2(-/-)-transferred mice with or without colitis showed phenotypes of memory cells, but only CD4(+) LPLs from colitic mice showed IL-7R(high). In vitro stimulation by IL-7, but not by IL-15 and thymic stromal lymphopoietin, enhanced significant proliferative responses and survival of colitic CD4(+), but not normal CD4(+) LPLs. Importantly, in vivo administration of IL-7 mice accelerated the expansion of IL-7R(high) memory CD4(+) LPLs and thereby exacerbated chronic colitis in RAG-2(-/-) mice transferred with CD4(+) LPLs from colitic TCR-alpha(-/-) mice. Conversely, the administration of anti-IL-7R monoclonal antibody significantly inhibited the development of TCR-alpha(-/-) colitis with decreased expansion of CD4(+) LPLs. Collectively, the present data indicate that IL-7 is essential for the expansion of pathogenic memory CD4(+) T cells under pathological conditions. Therefore, therapeutic approaches targeting the IL-7R pathway may be feasible in the treatment of human inflammatory bowel disease.     

Fission yeast MO25 protein is localized at SPB and septum and is essential for cell morphogenesis

Cell morphogenesis is of fundamental significance in all eukaryotes for development, differentiation, and cell proliferation. In fission yeast, Drosophila Furry-like Mor2 plays an essential role in cell morphogenesis in concert with the NDR/Tricornered kinase Orb6. Mutations of these genes result in the loss of cell polarity. Here we show that the conserved proteins, MO25-like Pmo25, GC kinase Nak1, Mor2, and Orb6, constitute a morphogenesis network that is important for polarity control and cell separation. Intriguingly, Pmo25 was localized at the mitotic spindle pole bodies (SPBs) and then underwent translocation to the dividing medial region upon cytokinesis. Pmo25 formed a complex with Nak1 and was required for both the localization and kinase activity of Nak1. Pmo25 and Nak1 in turn were essential for Orb6 kinase activity. Further, the Pmo25 localization at the SPBs and the Nak1-Orb6 kinase activities during interphase were under the control of the Cdc7 and Sid1 kinases in the septation initiation network (SIN), suggesting a functional linkage between SIN and the network for cell morphogenesis/separation following cytokinesis.     

LET and ion species dependence for cell killing in normal human skin fibroblasts

We studied the LET and ion species dependence of the RBE for cell killing to clarify the differences in the biological effects caused by the differences in the track structure that result from the different energy depositions for different ions. Normal human skin fibroblasts were irradiated with heavy-ion beams such as carbon, neon, silicon and iron ions that were generated by the Heavy Ion Medical Accelerator in Chiba (HIMAC) at the National Institute of Radiological Science (NIRS) in Japan. Cell killing was measured as reproductive cell death using a colony formation assay. The RBE-LET curves were different for carbon ions and for the other ions. The curve for carbon ions increased steeply up to around 98 keV/microm. The RBE of carbon ions at 98 keV/microm was 4.07. In contrast, the curves for neon, silicon and iron ions had maximum peaks around 180 keV/microm, and the RBEs at the peak position ranged from 3.03 to 3.39. When the RBEs were plotted as a function of Z*2/beta2 (where Z* is the effective charge and beta is the relative velocity of the ion) instead of LET, the discrepancies between the RBE-LET curves for the different ion beams were reduced, but branching of the RBE-Z*2/beta2 curves still remained. When the inactivation cross section was plotted as a function of either LET or Z*2/beta2, it increased with increasing LET. However, the inactivation cross section was always smaller than the geometrical cross section. These results suggest that the differences in the energy deposition track structures of the different ion sources have an effect on cell killing.     

Mono-(2-ethylhexyl) phthalate (MEHP) induces testicular alterations in male guinea pigs at prepubertal stage

We have recently shown that MEHP induces spermatogenic cell apoptosis in guinea pigs at prepubertal stage in vitro. To evaluate the effects of MEHP on the testicular tissues of guinea pigs in vivo, we conducted this research work. Five weeks old male guinea pigs were used in this experiment. They received a single oral dose of 2000 mg/ml of MEHP in corn oil by gavage at a volume equal to 4 ml/kg. Control group received a similar volume of corn oil vehicle. Vehicle- and MEHP-treated guinea pigs were sacrificed at the interval of 3, 6, and 9 h, and the testicular tissues were processed for histopathological studies. Distinct histopathological changes were recognized in testes. Detachment and displacement of spermatogenic cells, thin seminiferous epithelia, vacuolization of Sertoli cells were prominent at 6 h after MEHP treatment. The lumina of the efferent ductules were frequently occupied with sloughed seminiferous epithelia from 6 to 9 h after MEHP treatment. Apoptotic spermatogenic cells appeared at 3 h in the control group. The incidence of apoptotic spermatogenic cells significantly increased (*p<0.05) from 3 to 9 h, and the maximal increase of apoptotic spermatogenic cells were observed at 9 h after MEHP treatment. Time-dependent increases of apoptotic spermatogenic cells was recognized throughout the experimental period. It may be suggested here that MEHP also induces spermatogenic cell apoptosis in guinea pigs in vivo and guinea pigs may be considered as a useful animal model for sensitivity test of the reproductive toxicity to some phthalate esters at their earlier stage in vivo.     

Aged garlic extract inhibits peroxynitrite-induced hemolysis

Nitric oxide (NO), which is synthesized by constitutive NO synthase (cNOS), plays important roles in physiological functions of the cardiovascular system. However, NO, which is synthesized by inducible NOS, is detrimental when it reacts with superoxide to form peroxynitrite. Peroxynitrite is recognized as a powerful oxidant, and results in vascular or tissue damage. We have previously reported that aged garlic extract (AGE) enhances NO production through cNOS stimulation. In the present study, we determined the effect of AGE, its fractions or constituents on peroxynitrite-induced hemolysis using rat erythrocytes. Incubation of rat erythrocytes with peroxynitrite (300 microM) for 30 min at 37 degrees C caused 4-fold hemolysis. AGE (0.14-0.57 %w/v) added to an erythrocyte suspension was found to reduce peroxynitrite-induced hemolysis in a concentration-dependent manner. Of the AGE fractions, a polar fraction and a low-molecular-weight fraction both suppressed the hemolysis to the same degree as that seen with AGE. S-allylcysteine, one of the major compounds in AGE, also reduced hemolysis at 1-10 mM dose-dependently. These data indicate that AGE and its compounds protect erythrocytes from membrane damage induced by peroxynitrite, suggesting that AGE could be useful for prevention of cardiovascular diseases associated with oxidative stress or dysfunction of NO production.     

Discovery of potent thiosemicarbazone inhibitors of rhodesain and cruzain

Herein we report the synthesis and evaluation of a series of thiosemicarbazones as potential inhibitors of cysteine proteases relevant to parasitic diseases. Derivatives of thiosemicarbazone 1 were discovered to be potent inhibitors of cruzain and rhodesain, crucial proteases in the life cycles of Trypanosoma cruzi and T. brucei rhodesiense, the organisms causing Chagas' disease and sleeping sickness. However, the entire series had only modest potency against falcipain-2, an essential protease for Plasmodium falciparum, the organism causing malaria. Among the active inhibitors, several potently inhibited proliferation of cultures of T. brucei. However, only modest activity was observed in inhibition of proliferation of T. cruzi or P. falciparum.     

Suppression of matrix metalloproteinase production in nasal fibroblasts by tranilast, an antiallergic agent, in vitro

Allergic rhinitis is an inflammatory disease characterized by nasal wall remodeling with intense infiltration of eosinophils and mast cells/basophils. Matrix metalloproteinases (MMPs), MMP-2 and MMP-9, are the major proteolytic enzymes that induce airway remodeling. These enzymes are also important in the migration of inflammatory cells through basement membrane components. We evaluated whether tranilast (TR) could inhibit MMP production from nasal fibroblasts in response to tumor necrosis factor-alpha (TNF-alpha) stimulation in vitro. Nasal fibroblasts (NF) were established from nasal polyp tissues taken from patients with allergic rhinitis. NF (2 x 10(5) cells/mL) were stimulated with TNF-alpha in the presence of various concentrations of TR. After 24 hours, the culture supernatants were obtained and assayed for MMP-2, MMP-9, TIMP-1, and TIMP-2 levels by ELISA. The influence of TR on mRNA expression of MMPs and TIMPs in cells cultured for 12 hours was also evaluated by RT-PCR. TR at more than 5 x 10(-5) M inhibited the production of MMP-2 and MMP-9 from NF in response to TNF-alpha stimulation, whereas TIMP-1 and TIMP-2 production was scarcely affected. TR also inhibited MMP mRNA expression in NF after TNF-alpha stimulation. The present data suggest that the attenuating effect of TR on MMP-2 and MMP-9 production from NF induced by inflammatory stimulation may underlie the therapeutic mode of action of the agent in patients with allergic diseases, including allergic rhinitis.