Membrane TNF confers protection to acute mycobacterial infection

Background

Tumour necrosis factor (TNF) is crucial for the control of mycobacterial infection as TNF deficient (KO) die rapidly of uncontrolled infection with necrotic pneumonia. Here we investigated the role of membrane TNF for host resistance in knock-in mice with a non-cleavable and regulated allele (mem-TNF).

Methods

C57BL/6, TNF KO and mem-TNF mice were infected with M. tuberculosis H37Rv (Mtb at 100 CFU by intranasal administration) and the survival, bacterial load, lung pathology and immunological parameters were investigated. Bone marrow and lymphocytes transfers were used to test the role of membrane TNF to confer resistance to TNF KO mice.

Results

While TNF-KO mice succumbed to infection within 4–5 weeks, mem-TNF mice recruited normally T cells and macrophages, developed mature granuloma in the lung and controlled acute Mtb infection. However, during the chronic phase of infection mem-TNF mice succumbed to disseminated infection with necrotic pneumonia at about 150 days. Reconstitution of irradiated TNF-KO mice with mem-TNF derived bone marrow cells, but not with lymphocytes, conferred host resistance to Mtb infection in TNF-KO mice.

Conclusion

Membrane expressed TNF is sufficient to allow cell-cell signalling and control of acute Mtb infection. Bone marrow cells, but not lymphocytes from mem-TNF mice confer resistance to infection in TNF-KO mice. Long-term infection control with chronic inflammation likely disrupting TNF mediated cell-cell signalling, additionally requires soluble TNF.     

The effects of TNF-alpha and inhibitors of arachidonic acid metabolism on human colon HT-29 cells depend on differentiation status

The level of differentiation could influence sensitivity of colonic epithelial cells to various stimuli. In our study, the effects of TNF-alpha, inhibitors of arachidonic acid (AA) metabolism (baicalein, BA; indomethacin, INDO; niflumic acid, NA; nordihydroguaiaretic acid, NDGA), and/or their combinations on undifferentiated or sodium butyrate (NaBt)-differentiated human colon adenocarcinoma HT-29 cells were compared. NaBt-treated cells became growth arrested (blocked in G0/G1 phase of the cell cycle), and showed down-regulated Bcl-xL and up-regulated Bak proteins and increased expression of cyclooxygenase-2 (COX-2) and 5-lipoxygenase (5-LOX). These cells were more perceptive to anti-proliferative and apoptotic effects of TNF-alpha. Both inhibitors of LOX (BA and NDGA) and COX (INDO and NA) in higher concentrations modulated cell cycle changes accompanying NaBt-induced differentiation and induced various level of cell death in undifferentiated and differentiated cells. Most important is our finding that TNF-alpha action on proliferation and cell death can be potentiated by co-treatment of cells with AA metabolism inhibitors, and that these effects were more significant in undifferentiated cells. TNF-alpha and INDO co-treatment was associated with accumulation of cells in G0/G1 cell cycle phase, increased reactive oxygen species production, and elevated caspase-3 activity. These results indicate the role of differentiation status in the sensitivity of HT-29 cells to the anti-proliferative and proapoptotic effects of TNF-alpha, AA metabolism inhibitors, and their combinations, and imply promising possibility for novel anti-cancer strategies.     

Necrosis predominates in the cell death of human colon adenocarcinoma HT-29 cells treated under variable conditions of photodynamic therapy with hypericin.

Photodynamic therapy (PDT) represents a new rapidly-developing anticancer approach based on administration of a non- or weakly-toxic photosensitizer and its activation with light of appropriate wavelength. Hypericin, one of the promising photosensitizers, is known to induce apoptosis with high efficiency in various cell line models. However, here we report the prevalence of necrosis accompanied by suppression of caspase-3 activation in colon adenocarcinoma HT-29 cells exposed to an extensive range of PDT doses evoked by variations in two variables -- hypericin concentration and light dose. Necrosis was the principal mode of cell death despite different PDT doses and the absence of anti-apoptotic Bcl-2 expression, even if the same condition induced caspase-3 activity at similar toxicity in HeLa cells. Introduction of Bcl-2 into HT-29 cells invoked caspase-3 activation, changed the Bcl-X(L) expression pattern, increased the apoptosis ratio with no effect on overall toxicity, and supported arrest in the G(2)/M-phase of cell cycle. Since it is known that Bcl-2 suppression in HT-29 is reversible and linked to the over-expression of mutated p53 and also considering our data, we suggest that the mutation in p53 and events linked to this feature may play a role in cell death signalling in HT-29 colon cancer cells.     

Scrotal circumference of two-year-old bulls of several beef breeds

Scrotal circumference (SC) was measured on 7,918 2-yr-old Angus, Charolais, horned and polled Herefords, Limousin, Shorthorn, and Simmental bulls presented to culling committees at six show/sales between 1977 and 1983. Only SC data from bulls within the age range of 24 +/- 4 mo were used. Scrotal circumference data were corrected across breeds for the effects of location-year and sire and were adjusted to a common bull age of 730 d. The adjusted mean SC (+/- SE) for 2-yr-old beef bulls was Simmental, 38.8 +/- 0.10 cm (n = 540); Aberdeen Angus, 37.2 +/- 0.09 cm (n = 629); Charolais, 36.3 +/- 0.09 cm (n = 499); horned Hereford, 36.1 +/- 0.03 cm (n = 3,769); polled Hereford, 35.6 +/- 0.04 cm (n = 2,170); Shorthorn, 34.9 +/- 0.11 cm (n = 231); and Limousin, 32.2 +/- 0.18 cm (n = 80). The authors' recommendations of minimum acceptable SC for 2-yr-old beef bulls are Simmental, 36.0 cm; Angus and Charolais, 35.0 cm; horned and polled Herefords and Shorthorn, 34.0 cm; and Limousin, 33.0 cm.     

Defensive insect symbiont leads to cascading extinctions and community collapse

Animals often engage in mutualistic associations with microorganisms that protect them from predation, parasitism or pathogen infection. Studies of these interactions in insects have mostly focussed on the direct effects of symbiont infection on natural enemies without studying community‐wide effects. Here, we explore the effect of a defensive symbiont on population dynamics and species extinctions in an experimental community composed of three aphid species and their associated specialist parasitoids. We found that introducing a bacterial symbiont with a protective (but not a non‐protective) phenotype into one aphid species led to it being able to escape from its natural enemy and increase in density. This changed the relative density of the three aphid species which resulted in the extinction of the two other parasitoid species. Our results show that defensive symbionts can cause extinction cascades in experimental communities and so may play a significant role in the stability of consumer‐herbivore communities in the field.     

β-Arrestin Promotes Wnt-induced Low Density Lipoprotein Receptor-related Protein 6 (Lrp6) Phosphorylation via Increased Membrane Recruitment of Amer1 Protein

β-Arrestin is a scaffold protein that regulates signal transduction by seven transmembrane-spanning receptors. Among other functions it is also critically required for Wnt/β-catenin signal transduction. In the present study we provide for the first time a mechanistic basis for the β-arrestin function in Wnt/β-catenin signaling. We demonstrate that β-arrestin is required for efficient Wnt3a-induced Lrp6 phosphorylation, a key event in downstream signaling. β-Arrestin regulates Lrp6 phosphorylation via a novel interaction with phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P₂)-binding protein Amer1/WTX/Fam123b. Amer1 has been shown very recently to bridge Wnt-induced and Dishevelled-associated PtdIns(4,5)P₂ production to the phosphorylation of Lrp6. Using fluorescence recovery after photobleaching we show here that β-arrestin is required for the Wnt3a-induced Amer1 membrane dynamics and downstream signaling. Finally, we show that β-arrestin interacts with PtdIns kinases PI4KIIα and PIP5KIβ. Importantly, cells lacking β-arrestin showed higher steady-state levels of the relevant PtdInsP and were unable to increase levels of these PtdInsP in response to Wnt3a. In summary, our data show that β-arrestins regulate Wnt3a-induced Lrp6 phosphorylation by the regulation of the membrane dynamics of Amer1. We propose that β-arrestins via their scaffolding function facilitate Amer1 interaction with PtdIns(4,5)P₂, which is produced locally upon Wnt3a stimulation by β-arrestin- and Dishevelled-associated kinases.     

Monthly day/night changes and seasonal daily rhythms of sexual steroids in Senegal sole (Solea senegalensis) under natural fluctuating or controlled environmental conditions

In this paper we attempted to investigate the existence of daily fluctuations on plasma sexual steroids (17beta-estradiol, E(2) and testosterone, T) in Senegal sole (Solea senegalensis) females. We described the monthly day/night concentrations and seasonal daily rhythms in animals reared under natural photo- and thermo-period. In addition, the influence of the natural annual fluctuation of the water temperature on the plasma concentration of these steroids was investigated, using one group of Senegal sole under a natural photoperiod, but with an attenuated thermal cycle (around 17-20 degrees C) for one year. Although no significant day/night differences were detected in monthly samplings, the existence of an annual rhythm of E(2) and T (p<0.01) with an acrophase in February was revealed by COSINOR analysis. Maximum values were reached in March for both steroids (6.1+/-1.7 ng mL(-1) at mid-dark, MD and 4.0+/-0.6 ng mL(-1) at mid-light, ML for E2 and 1.4+/-0.4 ng mL(-1) at MD and 0.8+/-0.1 ng mL(-1) at ML for T) in anticipation of the spawning season (May-June). As regards seasonal daily rhythms, the presence of daily oscillations was revealed. At the spring solstice (21st March) a daily rhythm was observed for both steroids (COSINOR, p<0.01), with an acrophase at 20:00 h (E(2)) and at 21:08 h (T). In summer, autumn and winter no daily rhythms were observed due to the low steroid levels at those seasons. When Senegal sole females were submitted to an attenuated annual thermal cycle, the steroid rhythm disappeared (there was no surge in spring, as in the control group) and these fish did not spawn, despite being subjected to natural photoperiod conditions. This result underlined the importance of the natural annual fluctuation of water temperature and photoperiod on the synchronization of the spawning season and on the onset of steroidogenesis.     

Screening of high-quality bread wheat dihaploid lines by the use of biochemical markers

This work was undertaken to evaluate the storage protein composition of 13 doubled haploid bread wheat (Triticum aestivum L.) lines in order to identify those carrying promising alleles. For this, 15–20 seeds per each doubled haploid line were used to determine alleles at the loci for high-molecular-weight glutenin subunits and gliadins. Acid polyacrylamide gel electrophoresis was applied to identify glianin alleles, whereas SDS electrophoresis was used in the case of high-molecular-weight glutenin subunits. The identification of doubled haploid lines (DHLs) with respect to their storage protein composition enabled the classification of the DHLs derived from the cross Acheloos × Vergina in four different classes. Furthermore, protein composition analysis revealed that DHLs derived from the F₁ Penios × Kavkaz were identical. In addition, these lines were found to carry 1BL/1RS translocation, which is associated with high yield potential and resistance to biotic and abiotic stress conditions. Finally, it was observed that, in two cases, one rare biotype of the parental varieties was involved in the crosses. Published in Russian in Fiziologiya Rastenii, 2006, Vol. 53, No. 3, pp. 444–448. The text was submitted by the authors in English.