SUMMARYA mouse model of pulmonary tuberculosis induced by the intratracheal instillation of live and virulent mycobacteria strain H37-Rv was used to examine the relationship of the histopathological findings with the local kinetics production and cellular distribution of tumour necrosis factor-a (TNF-a), interleukin-1a (IL-1a) and transforming growth factor-b ( TGF-b). The histopathological and immunological studies showed two phases of the disease: acute or early and chronic or advanced. The acute phase was characterized by inflammatory infiltrate in the alveolar-capillary interstitium, blood vessels and bronchial wall with formation of granulomas. During this acute phase, which lasted from 1 to 28 days, high percentages of TNF-a and IL-1a immunostained activated macrophages were observed principally in the interstium-intralveolar inflammatory infiltrate and in granulomas. Electron microscopy studies of these cells, showed extensive rough endoplasmic reticulum, numerous lysosomes and occasional mycobacteria. Double labelling with colloid gold showed that TNF-a and IL-1a were present in the same cells, but were confined to separate vacuoles near the Golgi area, and mixed in larger vacuoles near to cell membrane. The concentration of TNF-a and IL-1a as well as their respective mRNAs were elevated in the early phase, particularly at day 3 when the bacillary count decreased. A second peak was seen at days 14 and 21-28 when granulomas appeared and evolved to full maturation. In contrast, TGF-b production and numbers of immunoreactive cells were low in comparison with the advanced phase of the disease. The chronic phase was characterized by histopathological changes indicative of more severity (i.e. pneumonia, focal necrosis and extensive interstitial fibrosis) with a decrease in the TNF-a and IL-1a production that coincided with the highest level of TGF-b. The bacillary counts were highest as the macrophages became large, vacuolated foamy cells, and containing numerous bacilli with immunoreactivity to mycobacterial lipids and lipoarabinomannan ( LAM). These macrophages displayed poor and scarce TNF-a and IL-1a immunostaining but still strong immunoreactivity to TGF-b. These cytokine production kinetics and the spatial relationship between immunostained cells and lung lesions corroborate the important role of TNF-a and IL1a in the constitution of granulomas and immune protection during the early phase of the infection, and also suggest an important if not primary role for TGF-b in the immunopathogenesis of the advanced forms of pulmonary tuberculosis.

Coronavirus 19 Disease (COVID-19) originating in the province of Wuhan, China in 2019, is caused by the severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), whose infection in humans causes mild or severe clinical manifestations that mainly affect the respiratory system. So far, the COVID-19 has caused more than 2 million deaths worldwide. SARS-CoV-2 contains the Spike (S) glycoprotein on its surface, which is the main target for current vaccine development because antibodies directed against this protein can neutralize the infection. Companies and academic institutions have developed vaccines based on the S glycoprotein, as well as its antigenic domains and epitopes, which have been proven effective in generating neutralizing antibodies. However, the emergence of new SARS-CoV-2 variants could affect the effectiveness of vaccines. Here, we review the different types of vaccines designed and developed against SARS-CoV-2, placing emphasis on whether they are based on the complete S glycoprotein, its antigenic domains such as the receptor-binding domain (RBD) or short epitopes within the S glycoprotein. We also review and discuss the possible effectiveness of these vaccines against emerging SARS-CoV-2 variants.

pJVl is an 11 kb, high-copy-number conjugative Streptomyces phaeochromogenes plasmid that replicates by the rolling circle mechanism (RCR). Sequencing combined with functional analysis of deletion, insertion and frameshift mutations was used to characterize the genes involved in plasmid transfer and chromosome mobilization (Cma), the single-strand origin for RCR and an associated strong incompatibility (Sti) determinant. pJVl contains two essential transfer genes whose expression is regulated by an adjacent repressor gene with similarity to the GntR family of regulators. A consensus sequence specific for the helix-turn-helix motifs of repressor proteins of Streptomyces plasmids is proposed. Unregulated expression of the transfer genes by inactivation of the repressor is lethal. Three additional genes increase intramycelial plasmid spread resulting in pock formation but, unlike the essential transfer genes, are not required for Cma. The pJVl transfer genes and their regulatory region, but not the minimal replication region encoding the double-strand replication origin and replication protein, are similar in their sequence and arrangement to those of the Strepfomyces nigrifaciens plasmid pSN22, revealing a modular organization of Strepfomyces RCR plasmids.Keywords : Streptomyces, plasmid transfer, plasmid spread, modular organization, rollingcircle replication INTRODUCTION Members of the genus Streptomyes are high G + C contentGram-positive mycelial soil bacteria with a complex lifecycle (Chater, 1989(Chater, ,1993. Linear, circular and integrating plasmids, ranging in size from a few kilobases to several hundreds, are common in streptomycetes (reviewed by Hopwood et al., 1986 andHopwood &Kieser, 1993). Amongst these, the most studied are the 9-1 1 kb, circular Hagkge e t al., 1993b; Kataoka et al., 1991a, b All the Streptomyces plasmids used in this study (Table 1) were derived from pJV1, except for pB2 and its derivatives pB62, pB68 and pB70, which contain hybrid pJV1-pIJ101 ds origins of replication (Servin-Gonzilez, 1993). Fig. 1 shows maps of all Streptomyces plasmids. pB65, pB66 and pB67 were constructed by cloning the 3 kb EcoRI-EcoRV fragment from pB50 (the single EcoRV site is located inside the tsr gene) into EcoRI/ EcoRV-cut pBR322 (Bolivar e t al., 1977) and, after modifying the fragment, re-introducing it into pB50. Some DNA manipulations were carried out in E. coli using pB72, which was constructed by inserting the EcoRI-Hind111 polylinker fragment from pUCBM21 (Boehringer Mannheim) into PI 52925 (Janssen & Bibb, 1993) digested with EcoRI and HindIII, and then deleting the ApaI site in the polylinker with Klenow enzyme to restore blue-white colony selection; pB72, therefore has a modified pUCBM21 polylinker flanked by BgllI sites, and was specifically used to insert pJVl fragments into pB83 in both orientations (pB85, pB86, pB87, pB88; Fig. 1 and Table 1). pIJ486 (Ward e t al., 1986) was the source of the 1.1 kb Bch tsr fragment, pIJ702 (Katz et al., 1983) was the source of the 1.5 kb Bell mel f...

In the present study we have explored the role of calmodulin (CaM) and inositol 1,4,5-trisphosphate receptor (IP 3 R) in the communication process activated after the release of calcium from the endoplasmic reticulum (ER) and the activation of calcium influx via endogenous TRP1 channels from Chinese hamster ovary cells. Experiments using combined rapid confocal calcium and electrophysiology measurements uncovered a consistent delay of around 900 ms between the first detectable calcium released from the ER and the activation of the calcium current. This delay was evident with two different methods used to release calcium from the ER: either the blockade of the microsomal calcium ATPase with thapsigargin or activation of bradykinin receptors linked to the IP 3 cascade. Direct application of IP 3 or a peptide from the NH 2 -terminal region of the IP 3 R activated store operated calcium, reducing the delay period. Introduction of CaM into the cell via the patch pipette increased the delay period from 900 ؎ 100 ms to 10 ؎ 2.1 s (n ‫؍‬ 18). Furthermore, the use of selective CaM antagonists W7 and trifluoperazine maleate resulted in a substantial reduction of the delay period to 200 ؎ 100 ms with 5 M trifluoperazine maleate (n ‫؍‬ 16) and 150 ؎ 50 ms with 500 nM W7 (n ‫؍‬ 22). CaM reduced also the current density activated by thapsigargin or brandykinin to about 60% from control. The CaM antagonists did not affect significantly the current density. The results presented here are consistent with an antagonistic effect of IP 3 R and CaM for the activation of store operated calcium after depletion of the ER. The functional competition between the activating effect of IP 3 R and the inhibiting effect of CaM may modulate the delay period between the release of calcium from the ER and the activation of calcium influx observed in different cells, as well as the amount of current activated after depletion of the ER.In a wide variety of nonexcitable and in many excitable cells, activation of G-protein-coupled receptors initiates a linear sequence of events leading to depletion of intracellular calcium storage compartments (the endoplasmic reticulum, ER) 1 via the production of inositol 1,4,5-trisphosphate (IP 3 ) by phospholipase C, and the subsequent induction of calcium influx from the extracellular space (1).Depletion of the ER appears to be a prerequisite for the activation of calcium influx, because several experimental maneuvers that induce depletion of the ER, such as the introduction of IP 3 into the cell or blockade of the microsomal calcium ATPase with thapsigargin (TG) and other selective blockers, are equally effective activators of calcium influx (1).The activation of calcium influx after store depletion has been termed store-operated calcium entry (SOCE) and appears to be a well preserved mechanism from insects to humans (2). The finding that the transient receptor potential protein (TRP) from the Drosophila photoreceptor encodes a calcium-permeable channel activated after depletion of intracellular calcium stores...

We have found that sera from patients with early stages of Lyme disease contain predominant immunoglobulin M reactivity to a major 23-kDa protein (p23) from Borrelia burgdorfieri 2591 isolated in Connecticut. To characterize this immunodominant antigen, we cloned and sequenced p23 and found it to be 83% identical by nucleotide sequence and 75% identical by amino acid sequence to pC (recently renamed OspC), an abundantly expressed protein on the outer surface of PKo, a European strain of B. burgdorfieri (B. Wilske, V.

Oxidative stress is a biochemical state of imbalance in the production of reactive oxygen and nitrogen species and antioxidant defenses. It is involved in the physiopathology of degenerative and chronic neuronal disorders, such as epilepsy. Experimental evidence in humans and animals support the involvement of oxidative stress before and after seizures. In the past few years, research has increasingly focused on the molecular pathways of this process, such as that involving transcription factor nuclear factor E2-related factor 2 (Nrf2), which plays a central role in the regulation of antioxidant response elements (ARE) and modulates cellular redox status. The aim of this review is to present experimental evidence on the role of Nrf2 in this neurological disorder and to further determine the therapeutic impact of Nrf2 in epilepsy.

The involvement of inositol trisphosphate receptor (IP3R) in modulating store-operated calcium entry (SOCE) was established many years ago. Nevertheless, the molecular mechanism responsible for this observation has not been elucidated to this date. In the present study we show that IP3R associates to STIM1 upon depletion of the endoplasmic reticulum (ER) by activation of the inositol trisphosphate signaling cascade via G-protein coupled receptors. IP3R-STIM1 association results in enhanced STIM1 puncta formation and larger Orai-mediated whole-cell currents as well as increased calcium influx. Depleting the ER with a calcium ATPase inhibitor (thapsigargin, TG) does not induce IP3R-STIM1 association, indicating that this association requires an active IP3R. The IP3R-STIM1 association is only observed after IP3R activation, as evidenced by FRET experiments and co-immunoprecipitation assays. ER intraluminal calcium measurements using Mag-Fluo-4 showed enhanced calcium depletion when IP3R is overexpressed. A STIM1-GCaMP fusion protein indicates that STIM1 detects lower calcium concentrations near its EF-hand domain when IP3R is overexpressed when compared with the fluorescence reported by a GCaMP homogenously distributed in the ER lumen (ER-GCaMP). All these data together strongly suggest that activation of inositol trisphosphate signaling cascade induces the formation of the IP3R-STIM1 complex. The activated IP3R provides a reduced intraluminal calcium microenvironment near STIM1, resulting in enhanced activation of Orai currents and SOCE.

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