NADH Intraperitoneal Injection Prevents Lung Inflammation in a BALB/C Mice Model of Cigarette Smoke-Induced Chronic Obstructive Pulmonary Disease.

Cigarette smoke is one of the main factors in Chronic Obstructive Pulmonary Disease (COPD), a respiratory syndrome marked by persistent respiratory symptoms and increasing airway obstruction. Perturbed NAD+/NADH levels may play a role in various diseases, including lung disorders like COPD. In our study, we investigated the preventive effect of NADH supplementation in an experimental model of COPD induced by cigarette smoke extract (CSE). N = 64 mice randomly distributed in eight groups were injected with NADH (two doses of 100 mg/kg or 200 mg/kg) or dexamethasone (2 mg/kg) before being exposed to CSE for up to 9 weeks. Additionally, NADH supplementation preserved lung antioxidant defenses by preventing the functional loss of key enzymes such as superoxide dismutase (SOD), glutathione peroxidase (GPX), catalase, and the expression levels of glutathione (GSH) (n = 4, p < 0.001). It also reduced oxidative damage markers, such as malondialdehyde (MDA) and nitrites (n = 4, p < 0.001). A marked increase in tissue myeloperoxidase activity was assessed (MPO), confirming neutrophils implication in the inflammatory process. The latter was significantly ameliorated in the NADH-treated groups (p < 0.001). Finally, NADH prevented the CSE-induced secretion of cytokines such as Tumor Necrosis Factor alpha (TNF-α), IL-17, and IFN-y (n = 4, p < 0.001). Our study shows, for the first time, the clinical potential of NADH supplementation in preventing key features of COPD via its unique anti-inflammatory and antioxidant properties.

NADH intraperitoneal injection prevents massive pancreatic beta cell destruction in a streptozotocin-induced diabetes in rats.

Diabetes mellitus is a chronic metabolic disease characterized by persistent hyperglycemia, revealing a decrease in insulin efficiency. The sustained glucotoxic pancreatic microenvironment increases reactive oxygen species generation, resulting in chronic oxidative stress responsible for massive DNA damage. This triggers PARP-1 activation with both NAD+ and ATP depletion, affecting drastically pancreatic beta cells' energy storage and leading to their dysfunction and death. The aim of the present study is to highlight the main histological changes observed in pancreatic islets pre-treated with a unique NADH intraperitoneal injection in a streptozotocin-(STZ)-induced diabetes model. In order to adjust NADH doses, a preliminary study with three different doses, 500 mg/kg, 300 mg/kg, and 150 mg/kg, respectively, was conducted. Subsequently, and on the basis of the results of the aforementioned study, Wistar rats were randomly divided into four groups: non-diabetic control group, diabetics (STZ 45 mg/kg), NADH-treated group (150 mg/kg) 15 min before STZ administration, and NADH-treated group (150 mg/kg) 15 min after STZ administration. The effect of NADH was assessed by blood glucose level, TUNEL staining, histo-morphological analysis, and immunohistochemistry. The optimum protective dose of NADH was 150 mg/kg. NADH effectively decreased hyperglycemia and reduced diabetes induced by STZ. Histologically, NADH pre-treatment revealed a decrease in beta cell death favoring apoptosis over necrosis and therefore preventing inflammation with further beta cell destruction. Our data clearly demonstrate that NADH prior or post-treatment could effectively prevent the deleterious loss of beta cell mass in STZ-induced diabetes in rats and preserve the normal pancreatic islet's function.

First serological evidence of MERS-CoV in dromedary camels from Algeria.

Middle East Respiratory Syndrome (MERS) is a zoonotic disease. Dromedary camel is responsible of its transmission to humans. Accordingly, several human cases have been reported worldwide with a high mortality rate. In Algeria, no data reported on MERS prevalence in camels. This is a first seroprevalence study MERS-CoV in Algerian dromedaries. A total of 87 camel blood samples from EL -MENIAA and Ghardaia, were analyzed by anti-MERS-CoV IgG ELISA camel. The seroprevalence was 64 % and it significantly increases with age. Larger serological and molecular screening is needed to precisely determine the rate of MERS active circulation among Algerian dromedary population.

Pseudochrobactrum algeriensis sp. nov., isolated from lymph nodes of Algerian cattle.

Three Gram-negative, rod-shaped, oxidase-positive, non-spore-forming, non-motile strains (C130915_07T, C150915_16 and C150915_17) were isolated from lymph nodes of Algerian cows. On the basis of 16S rRNA gene and whole genome similarities, the isolates were almost identical and clearly grouped in the genus Pseudochrobactrum. This allocation was confirmed by the analysis of fatty acids (C19:cyclo, C18 : 1, C18 : 0, C16 : 1 and C16 : 0) and of polar lipids (major components: phosphatidylethanolamine, ornithine-lipids, phosphatidylglycerol, cardiolipin and phosphatidylcholine, plus moderate amounts of phosphatidylmonomethylethanolamine, phosphatidyldimethylethanolamine and other aminolipids). Genomic, physiological and biochemical data differentiated these isolates from previously described Pseudochrobactrum species in DNA relatedness, carbon assimilation pattern and growth temperature range. Thus, these organisms represent a novel species of the genus Pseudochrobactrum, for which the name Pseudochrobactrum algeriensis sp. nov. is proposed (type strain C130915_07T=CECT30232T=LMG 32378T).

First report on Cryptosporidium parvum, Escherichia coli K99, rotavirus and coronavirus in neonatal lambs from north-center region, Algeria.

The etiology of neonatal diarrhea is multifactorial and remains one of the greatest health problems in sheep livestock farming. Faecal samples from 559 neonatal lambs aged less than 30 days from 30 sheepfolds located in the north-center region of Algeria were screened with pathogen-specific antigen ELISA for Cryptosporidium parvum, Escherichia coli K99, rotavirus, and coronavirus. Of the 559 lambs, 312 (58.81 %), 155 (27.72 %), 72 (12.88 %) and 20 (3.57 %) were positives for C. parvum, E. coli K99, rotavirus and coronavirus antigens, respectively. The prevalence of C. parvum was the highest (p < 0.0001). C. parvum, E. coli K99, rotavirus and coronavirus were observed in 23 (76.66 %), 17 (56.66 %), 9 (30 %) and 3 (10 %) sheepfolds, respectively. Compared to age, the prevalence of C. parvum was highest during the second and third week of age (p < 0.001). In contrast, other pathogens were found to be more frequent in lambs aged ≤7 days (p < 0.001). The number of lambs with diarrhea was 280 (50.09 %) of which 280 (100 %), 127 (45.35 %), 52 (18.57 %) and 10 (3.57 %) were found to be infected with C. parvum, E. coli K99, rotavirus and coronavirus, respectively (p < 0.0001). In various combinations, mixed infections were detected only with C. parvum. This is the first report of C. parvum, E. coli K99, rotavirus, and coronavirus in ≤30-days old neonatal lambs in Algeria. Special attention should be given to the first colostrum feeding, hygiene of the farm, prevention and control measures for a better prevention of neonatal diarrhea in lambs.

GTS-21, an α7nAChR agonist, suppressed the production of key inflammatory mediators by PBMCs that are elevated in COPD patients and associated with impaired lung function.

Chronic obstructive pulmonary disease (COPD) is a lung inflammatory disease characterized by progressive airflow limitation, chronic respiratory symptoms and frequent exacerbations. There is an unmet need to identify novel therapeutic alternatives beside bronchodilators that prevent disease progression. Levels of both Nitric Oxide (NO) and IL-6 were significantly increased in the plasma of patients in the exacerbation phase (ECOPD, n = 13) when compared to patients in the stable phase (SCOPD, n = 38). Levels of both NO and IL-6 were also found to inversely correlate with impaired lung function (%FEV1 predicted). In addition, there was a strong positive correlation between levels of IL-6 and NO found in the plasma of patients and those spontaneously produced by their peripheral blood mononuclear cells (PBMCs), identifying these cells as a major source of these key inflammatory mediators in COPD. GTS-21, an agonist for the alpha 7 nicotinic receptors (α7nAChR), was found to exert immune-modulatory actions in PBMCs of COPD patients by suppressing the production of IL-6 and NO. This study provides the first evidence supporting the therapeutic potential of α7nAChR agonists in COPD due to their ability to suppress the production of key inflammatory markers associated with disease severity.

Vaccination with UV-inactivated nodavirus partly protects European sea bass against infection, while inducing few changes in immunity.

Developing viral vaccines through the ultraviolet (UV) inactivation of virus is promising technique since it is straightforward and economically affordable, while the resulting viruses are capable of eliciting an adequate antiviral immune response. Nodavirus (NNV) is a devastating virus that mainly affects European sea bass juveniles and larvae, causing serious economic losses in Mediterranean aquaculture. In this work, a potential vaccine consisting on UV-inactivated NNV (iNNV) was generated and administered to healthy juveniles of European sea bass to elucidate whether it triggers the immune response and improves their survival upon challenge. First, iNNV failed to replicate in cell cultures and its intraperitoneal administration to sea bass juveniles also failed to produce fish mortality and induction of the type I interferon (IFN) pathway, indicating that the NNV was efficiently inactivated. By contrast, iNNV administration induced significant serum non-specific antimicrobial activity as well as a specific antiviral activity and immunoglobulin M (IgM) titres against NNV. Interestingly, few changes were observed at transcriptional level in genes related to either innate or adaptive immunity, suggesting that iNNV could be modulating the immune response at protein or functional level. In addition, the iNNV vaccinated group showed improved survival, reaching a relative survival percentage of 57.9%. Moreover, challenged fish that had been vaccinated presented increased serum antibacterial, antiviral and IgM titres, as well as the higher transcription of mhc1a, ifn, isg15 and cd8a genes in brain, while in the head-kidney the transcription of mhc1a, mhc2b and cd8a was down-regulated and mx, isg15 and tcrb was up-regulated. Although the UV-inactivated vaccine against NNV showed promising results, more effort should be addressed to improving this prophylactic method by increasing our understanding of its action mechanisms, thus enabling the mortality rate of NNV to be further reduced.

The characterization of Brucella strains isolated from cattle in Algeria reveals the existence of a B. abortus lineage distinct from European and Sub-Saharan Africa strains.

Brucellosis is a zoonosis caused by bacteria of the genus Brucella that causes important economic losses and human suffering worldwide. Brucellosis control requires an understanding of the Brucella species circulating in livestock and humans and, although prevalent in African countries of the Mediterranean basin, data for this area are mostly restricted to isolates obtained from humans and small ruminants. Here, we report the characterization of twenty-four Brucella strains isolated from Algerian cattle. Bruce-ladder multiplex PCR and conventional biotyping showed that Algerian cattle are infected mostly by B. abortus biovar 3, and to less extent by B. abortus biovar 1 and B. melitensis biovar 3. Extended AMOS-ERY PCR showed that all Algerian B. abortus biovar 3 strains were of the subgroup 3b. Although by multi locus variable number of tandem repeats analysis (MLVA) most isolates were closer to the European counterparts, five strains displayed characteristics distinct from the European isolates and those of countries across the Sahara, including three repetitions of marker Bruce55. These five strains, plus an earlier isolate from an Algerian human patient, may represent a lineage close to clades previously described in Africa. These data provide the basis for additional molecular epidemiology studies in northern Africa and indicate that further bacteriological and molecular investigations are necessary for a complete understanding of the epidemiology of cattle brucellosis in countries north and south of the Sahara.

Cell-contact dependent inhibition of monocytes by airway epithelial cells and reversion by infection with Respiratory Syncytial Virus.

Airway epithelial cells (AEC) are the first line of defense against airborne infectious microbes and play an important role in regulating the local immune response. However, the interplay of epithelial cells and professional immune cells during both homeostasis and infection has only been partially studied. The present study was performed to determine how bronchial epithelial cells affect the activation of monocytes. Under healthy conditions, AECs were shown to inhibit reactivity of monocytes. We hypothesized that upon infection, monocytes might be released from inhibition by AECs. We report that direct contact of monocytes with unstimulated BEAS2B epithelial cells results in inhibition of TNF secretion by activated monocytes. In addition to the known soluble modulators, we show that cell contacts between epithelial cells and monocytes or macrophages also contribute to homeostatic inhibitory actions. We find AECs to express the inhibitory molecule PD-L1 and blockade of PD-L1 results in increased secretion of pro-inflammatory cytokines from monocytes. Contrary to the inhibitory activities during homeostasis, epithelial cells infected with Respiratory Syncitial Virus (RSV) induce a significant release of inhibition. However, release of inhibition was not due to modulation of PD-L1 expression in AECs. We conclude that airway epithelial cells control the reactivity of monocytes through direct and indirect interactions; however tonic inhibition can be reverted upon stimulation of AECs with RSV and thereof derived molecular patterns. The study confirms the important role of airway epithelial cells for local immune reactions.

Innate humoral immune parameters in Tilapia zillii under acute stress by low temperature and crowding.

Redbelly tilapia (Tilapia zillii; Gervais, 1848) is one of the most valuable freshwater species in North Africa representing an important part of the continental production, especially in brackish lakes. In Algeria, T. zillii is distributed in several lakes and tributaries of some rivers in the south. Though some attempts are in progress to culture this species, many investigations covering its biology and farm management are still needed. In this sense, this is the first study attempting to evaluate some of the T. zillii immune parameters and valuable data to assess their health and well-being status. Thus, we have determined the levels of serum peroxidases as well as the alternative complement, antiprotease and bactericidal activities. Furthermore, we have also evaluated the potential impact of two acute stress factors, commonly found in fish farms, in these parameters. Although it was assessed that fish exposed to low temperatures or crowding were stressed, as indicated by their increased serum levels of cortisol and glucose, both acute stressors failed to significantly affect serum peroxidases as well as antiprotease and complement activities. However, the bactericidal activity was reduced in general but only in those exposed to crowding reached statistical significance. Further studies are needed to characterise the immune response in T. zillii as well as the effects that farming stresses may produce.

ICAD deficiency in human colon cancer and predisposition to colon tumorigenesis: linkage to apoptosis resistance and genomic instability.

We previously showed that DNA fragmentation factor, which comprises a caspase-3-activated DNase (CAD) and its inhibitor (ICAD), may influence the rate of cell death by generating PARP-1-activating DNA breaks. Here we tested the hypothesis that ICAD-deficient colon epithelial cells exhibiting resistance to death stimuli may accumulate additional genetic modifications, leading to a tumorigenic phenotype. We show that ICAD deficiency may be associated with colon malignancy in humans. Indeed, an examination of ICAD expression using immunohistochemistry in an array of both colon cancer and normal tissues revealed that ICAD expression levels were severely compromised in the cancerous tissues. Upon DNA damage caused by a low dose of irradiation, ICAD cells acquire a tumorigenic phenotype. Colon epithelial cells derived from ICAD mice showed a significant resistance to death induced by the colon carcinogen dimethylhydrazine in vitro and in mice. Such resistance was associated with a decrease in PARP-1 activation. In an animal model of dimethylhydrazine-induced colon tumorigenesis, ICAD(-/-) mice developed significantly higher numbers of tumors with markedly larger sizes than the wild-type counterparts. Interestingly, the phenotype of the ICAD(-/-) mice was not associated with a significant increase in the precancerous aberrant crypt foci suggesting a potential link to tumor progression rather than initiation. More importantly, ICAD deficiency was associated with severe genomic instability as assessed by array comparative genomic hybridization. Such genomic instability consisted most prominently of amplifications but with sizable deletions as compared to the wild-type counterparts affecting several cancer-related genes including RAF-1, GSN, LMO3, and Fzd6 independently of p53. Altogether, our results present a viable case for the involvement of ICAD deficiency in colon carcinogenesis and show that apoptosis and genomic instability may comprise the means by which such deficiency may contribute to the process of increasing susceptibility to carcinogen-induced tumorigenesis.

Cytotoxic responses to BLV tax oncoprotein do not prevent leukemogenesis in sheep.

Delta retrovirus-mediated leukemogenesis is dependent on the oncogenic potential of Tax. It is not clear, however, whether Tax-specific immune responses play a role in leukemia onset and progression. Using the BLV-associated leukemia model in sheep, we found that Tax-specific cytotoxic responses induced by DNA immunization or viral infection of naïve animals were not predictive of disease outcome and did not prevent tumor development. Furthermore, provirus and tax may be absent from blood for extended periods, emphasizing the relevance of surveying other compartments during chronic lymphoproliferative disorders. Our results support the conclusion that Tax-specific cytotoxic responses, even during the initial phase of infection, are not sufficient to prevent leukemogenesis.

Intranasal immunization of mice with a formalin-inactivated bovine respiratory syncytial virus vaccine co-formulated with CpG oligodeoxynucleotides and polyphosphazenes results in enhanced protection.

As respiratory syncytial virus (RSV) targets the mucosal surfaces of the respiratory tract, induction of both systemic and mucosal immunity will be critical for optimal protection. In this study, the ability of an intranasally delivered, formalin-inactivated bovine RSV (FI-BRSV) vaccine co-formulated with CpG oligodeoxynucleotides (ODN) and polyphosphazenes (PP) to induce systemic and mucosal immunity, as well as protection from BRSV challenge, was evaluated. Intranasal immunization of mice with FI-BRSV formulated with CpG ODN and PP resulted in both humoral and cell-mediated immunity, characterized by enhanced production of BRSV-specific serum IgG, as well as increased gamma interferon and decreased interleukin-5 production by in vitro-restimulated splenocytes. These mice also developed mucosal immune responses, as was evident from increased production of BRSV-specific IgG and IgA in lung-fragment cultures. Indeed, the increases in serum and mucosal IgG, and in particular mucosal IgA and virus-neutralizing antibodies, were the most critical differences observed between FI-BRSV formulated with both CpG ODN and PP in comparison to formulations with CpG ODN, non-CpG ODN or PP individually. Finally, FI-BRSV/CpG/PP was the only formulation that resulted in a significant reduction in viral replication upon BRSV challenge. Co-formulation of CpG ODN and PP is a promising new vaccine platform technology that may have applications in mucosal immunization in humans.

Intrinsic resistance to apoptosis of colon epithelial cells is a potential determining factor in the susceptibility of the A/J mouse strain to dimethylhydrazine-induced colon tumorigenesis.

Alterations in the delicate balance between cell proliferation and cell death disrupt colon homeostasis and serve as determining factors in colon tumorigenesis. The two mouse strains, AKR/J (resistant) and A/J (susceptible), have been widely used as models for dimethylhydrazine-induced colon tumorigenesis. This study examined whether the differential susceptibilities of the two mouse strains to the tumorigenic effect of dimethylhydrazine were associated with intrinsic differences in the apoptotic machinery of the colon epithelial cells. While acute exposure to dimethylhydrazine caused massive apoptosis of colon epithelial cells in AKR/J mice, the effect was considerably less in A/J mice. Apoptosis in AKR/J mice occurred not only in the luminal side of the mucosa but also deep in the colonic crypts. In addition, this apoptosis appeared to involve caspase-3. The increased sensitivity of AKR/J to dimethylhydrazine was associated with a persistent expression of tumor necrosis factor (TNF) but not of its receptors. After establishing a new method for isolating primary colon epithelial cells, we determined that cells derived from A/J mice were substantially more resistant to apoptosis in response to dimethylhydrazine or to a combination of TNF, cyclohexamide, and butyrate compared to cells from AKR/J mice. These results strongly suggest that a higher intrinsic resistance to apoptosis of colon epithelial cells may be an important determinant of predisposition to colon tumorigenesis in the A/J mouse strain.

Poly(ADP-ribose) polymerase-1 inhibition prevents eosinophil recruitment by modulating Th2 cytokines in a murine model of allergic airway inflammation: a potential specific effect on IL-5.

We recently used a murine model of allergic airway inflammation to show that poly(ADP-ribose) polymerase-1 (PARP-1) plays an important role in the pathogenesis of asthma-related lung inflammation. In this study, we show that PARP-1 inhibition, by a novel inhibitor (TIQ-A) or by gene deletion, prevented eosinophilic infiltration into the airways of OVA-challenged mice. Such impairment of eosinophil recruitment appeared to take place after IgE production. OVA challenge of wild-type mice resulted in a significant increase in IL-4, IL-5, IL-10, IL-13, and GM-CSF secretions. Although IL-4 production was moderately affected in OVA-challenged PARP-1(-/-) mice, the production of IL-5, IL-10, IL-13, and GM-CSF was completely inhibited in ex vivo OVA-challenged lung cells derived from these animals. A single TIQ-A injection before OVA challenge in wild-type mice mimicked the latter effects. The marked effect PARP-1 inhibition exerted on mucus production corroborated the effects observed on the Th2 response. Although PARP-1 inhibition by gene knockout increased the production of the Th1 cytokines IL-2 and IL-12, the inhibition by TIQ-A exerted no effect on these two cytokines. The failure of lung cells derived from OVA-challenged PARP-1(-/-) mice to synthesize GM-CSF, a key cytokine in eosinophil recruitment, was reestablished by replenishment of IL-5. Furthermore, intranasal administration of IL-5 restored the impairment of eosinophil recruitment and mucus production in OVA-challenged PARP-1(-/-) mice. The replenishment of either IL-4 or IgE, however, did not result in such phenotype reversals. Altogether, these results suggest that PARP-1 plays a critical role in eosinophil recruitment by specifically regulating the cascade leading to IL-5 production.

Formulation with CpG oligodeoxynucleotides increases cellular immunity and protection induced by vaccination of calves with formalin-inactivated bovine respiratory syncytial virus.

Vaccination of calves with formalin-inactivated bovine respiratory syncytial virus (FI-BRSV) induces low levels of cellular immunity that may not be protective. Since inactivated and subunit vaccines formulated with CpG oligodeoxynucleotides (ODNs) have been shown to induce cellular immune responses, we studied the ability of a FI-BRSV vaccine formulated with CpG ODN to elicit cellular immunity against BRSV. Neonatal calves were immunized with FI-BRSV, FI-BRSV formulated with CpG ODN or medium and challenged with BRSV after two immunizations. Calves vaccinated with FI-BRSV formulated with CpG ODN developed increased numbers of IFN-gamma secreting cells in the peripheral blood and broncho-tracheal lymph nodes and enhanced BRSV-specific serum IgG2 in comparison to FI-BRSV immunized animals. Calves that received the FI-BRSV vaccine formulated with CpG ODN also experienced a reduction in the amount of BRSV in the lung tissue. Based on these observations, CpG ODN appears to be a suitable candidate adjuvant for inactivated BRSV vaccines.

Correlation between decreased sensitivity of the Daudi lymphoma cells to VP-16-induced apoptosis and deficiency in DNAS1L3 expression.

A functional relationship between the apoptotic endonuclease DNAS1L3 and the chemotherapeutic drug VP-16 was established. The lymphoma cell line, Daudi, exhibited a significant resistance to VP-16 treatment in comparison to the lymphoma/leukemia cell line, U-937. While U-937 cells degraded their DNA into internucleosomal fragments, Daudi cells failed to undergo such fragmentation in response to the drug. Activation of both caspase-3 and DNA fragmentation factor was not sufficient to trigger internucleosomal DNA fragmentation in Daudi cells. No correlation was found between expression levels of topoisomerase-II, Pgp, Bcl-2, Bax, or Bad and decreased sensitivity of Daudi cells to VP-16. Daudi cells failed to express DNAS1L3 and ectopic expression of this protein significantly sensitized the cells to VP-16. An enhancement of caspase-3 activity and collapse of mitochondrial membrane potential underlie DNAS1L3-mediated sensitization of Daudi cells to VP-16, which may be a direct result of DNAS1L3-mediated increase in PARP-1-activating DNA breaks after VP-16 treatment. Our results suggest that DNAS1L3 plays an active role in lymphoma cell sensitization to VP-16 and that its deficiency may constitute a novel mechanism of drug resistance in these cells.