MUC22, HLA-A, and HLA-DOB variants and COVID-19 in resilient super-agers from Brazil.

Background: Although aging correlates with a worse prognosis for Covid-19, super elderly still unvaccinated individuals presenting mild or no symptoms have been reported worldwide. Most of the reported genetic variants responsible for increased disease susceptibility are associated with immune response, involving type I IFN immunity and modulation; HLA cluster genes; inflammasome activation; genes of interleukins; and chemokines receptors. On the other hand, little is known about the resistance mechanisms against SARS-CoV-2 infection. Here, we addressed polymorphisms in the MHC region associated with Covid-19 outcome in super elderly resilient patients as compared to younger patients with a severe outcome. Methods: SARS-CoV-2 infection was confirmed by RT-PCR test. Aiming to identify candidate genes associated with host resistance, we investigated 87 individuals older than 90 years who recovered from Covid-19 with mild symptoms or who remained asymptomatic following positive test for SARS-CoV-2 as compared to 55 individuals younger than 60 years who had a severe disease or died due to Covid-19, as well as to the general elderly population from the same city. Whole-exome sequencing and an in-depth analysis of the MHC region was performed. All samples were collected in early 2020 and before the local vaccination programs started. Results: We found that the resilient super elderly group displayed a higher frequency of some missense variants in the MUC22 gene (a member of the mucins' family) as one of the strongest signals in the MHC region as compared to the severe Covid-19 group and the general elderly control population. For example, the missense variant rs62399430 at MUC22 is two times more frequent among the resilient super elderly (p = 0.00002, OR = 2.24). Conclusion: Since the pro-inflammatory basal state in the elderly may enhance the susceptibility to severe Covid-19, we hypothesized that MUC22 might play an important protective role against severe Covid-19, by reducing overactive immune responses in the senior population.

MHC Variants Associated With Symptomatic Versus Asymptomatic SARS-CoV-2 Infection in Highly Exposed Individuals.

Despite the high number of individuals infected by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) who develop coronavirus disease 2019 (COVID-19) symptoms worldwide, many exposed individuals remain asymptomatic and/or uninfected and seronegative. This could be explained by a combination of environmental (exposure), immunological (previous infection), epigenetic, and genetic factors. Aiming to identify genetic factors involved in immune response in symptomatic COVID-19 as compared to asymptomatic exposed individuals, we analyzed 83 Brazilian couples where one individual was infected and symptomatic while the partner remained asymptomatic and serum-negative for at least 6 months despite sharing the same bedroom during the infection. We refer to these as "discordant couples". We performed whole-exome sequencing followed by a state-of-the-art method to call genotypes and haplotypes across the highly polymorphic major histocompatibility complex (MHC) region. The discordant partners had comparable ages and genetic ancestry, but women were overrepresented (65%) in the asymptomatic group. In the antigen-presentation pathway, we observed an association between HLA-DRB1 alleles encoding Lys at residue 71 (mostly DRB1*03:01 and DRB1*04:01) and DOB*01:02 with symptomatic infections and HLA-A alleles encoding 144Q/151R with asymptomatic seronegative women. Among the genes related to immune modulation, we detected variants in MICA and MICB associated with symptomatic infections. These variants are related to higher expression of soluble MICA and low expression of MICB. Thus, quantitative differences in these molecules that modulate natural killer (NK) activity could contribute to susceptibility to COVID-19 by downregulating NK cell cytotoxic activity in infected individuals but not in the asymptomatic partners.

KIR2DL4 genetic diversity in a Brazilian population sample: implications for transcription regulation and protein diversity in samples with different ancestry backgrounds.

KIR2DL4 is an important immune modulator expressed in natural killer cells; HLA-G is its main ligand. We have characterized the KIR2DL4 genetic diversity by considering the promoter, all exons, and all introns in a highly admixed Brazilian population sample and by using massively parallel sequencing. We introduce a molecular method to amplify and to sequence the complete KIR2DL4 gene. To avoid the mapping bias and genotype errors commonly observed in gene families, we have developed and validated a bioinformatic pipeline designed to minimize these errors and applied it to survey the variability of 220 individuals from the State of São Paulo, southeastern Brazil. We have also compared the KIR2DL4 genetic diversity in the Brazilian cohort with the diversity previously reported by the 1000Genomes consortium. KIR2DL4 presents high linkage disequilibrium throughout the gene, with coding sequences associated with specific promoters. There are few but divergent promoter haplotypes. We have also detected many new KIR2DL4 sequences, all bearing nucleotide exchanges in introns and encoding previously described proteins. Exons 3 and 4, which encode the external domains, are the most variable. The ancestry background influences the KIR2DL4 allele frequencies and must be considered for association studies regarding KIR2DL4.

Elevated circulatingadenosine deaminase activity in women with preeclampsia: association with pro-inflammatory cytokine production and uric acid levels.

OBJECTIVE: Preeclampsia is a specific disorder of human pregnancy that is associated with hyperuricemia and higher levels of pro-inflammatory cytokines. Adenosine deaminase (ADA) is an enzyme present in all human tissues, and is considered an indicator of cellular inflammation. In the present study we assess whether adenosine deaminase (ADA) activity is altered in women with preeclampsia (PE) and contributes to elevated levels of uric acid and pro-inflammatory cytokine production. STUDY DESIGN: The population studied consisted of 60 women with PE, 30 normotensive pregnant women (NT) and 20 non-pregnant women (NP). Uric acid concentration and ADA activity were determined in the serum. Peripheral blood mononuclear cells (PBMCs) were isolated and evaluated for intracellular nuclear transcription factor kappa B (NF-κB) levels and for endogenous tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß) production. The data were evaluated with parametric or non-parametric tests with significance set at P<0.05. RESULTS: ADA levels were higher in the PE group compared with the NT and NP groups (P<0.001). A positive correlation between ADA and uric acid levels was identified in women with PE (P<0.001). Endogenous production of IL-1ß and TNF-α, as well as intracellular NF-κB levels, were higher in PBMCs from the PE group than from NT and NP women (P<0.01) and correlated with the ADA concentration in preeclamptic women (P<0.01). CONCLUSION: An elevation in ADA activity in women with PE may contribute to their increased levels of uric acid and pro-inflammatory immune activity.

Monocytes from pregnant women with pre-eclampsia are polarized to a M1 phenotype.

PROBLEM: This study evaluated whether the monocyte inflammatory state in pre-eclampsia (PE) might be associated with polarization to either M1 classically or M2 alternatively activated monocyte subsets. METHOD OF STUDY: Eighty-five women with (PE) and 52 normotensive (NT) pregnant women matched for gestational age were included. Expression of surface receptors characteristic of M1, such as Toll-like receptor (TLR)2, TLR4, and CD64, or M2, such as CD163 and CD206 monocyte subsets were evaluated in peripheral blood monocytes by flow cytometry. Tumour necrosis factor-alpha (TNF-α), interleukin-(IL)-12p40, IL-12p70, and IL-10 were evaluated in the supernatant of monocyte cultures by ELISA. RESULTS: Expression of TLR4 and CD64 by monocytes from pre-eclamptic women was significantly higher, while the expression of CD163 and CD206 expression was significantly lower compared with NT pregnant women. Endogenous production of TNF-α, IL-12p40, and IL-12p70 by monocytes was increased, while synthesis of IL-10 was lower in women with PE than in NT pregnant women. CONCLUSIONS: Monocytes from women with PE are classically activated, producing higher levels of pro-inflammatory cytokines, and express surface receptors characteristic of the M1 subset. These results provide evidence that the systemic inflammatory environment in PE may differentiate and polarize these cells to the M1 phenotype.

High levels of heat shock protein 70 are associated with pro-inflammatory cytokines and may differentiate early- from late-onset preeclampsia.

Preeclampsia (PE), a specific syndrome of pregnancy, can be classified into early and late onset, depending on whether clinical manifestations occur before or after 34 weeks' gestation. We determined whether plasma concentrations of Hsp60 and Hsp70 were related to circulating cytokine levels, as well as kidney and liver functions, in early- and late-onset PE. Two hundred and thirty-seven preeclamptic women (95 with early- and 142 with late-onset PE) were evaluated. Plasma levels of Hsp60, Hsp70, and their specific antibodies, tumor necrosis factor-alpha (TNF-α), interleukin (IL)-1, IL-10, IL-12, and soluble TNF-α-receptor I (sTNFRI) concentrations, were determined by enzyme-linked immunosorbent assay (ELISA). Concentrations of Hsp70, TNF-α, IL-1ß, IL-12, and sTNFRI were significantly elevated in patients with early-onset PE compared with women with late-onset PE; IL-10 levels were significantly lower in the early-onset PE group. Concentrations of urea, uric acid, proteinuria, glutamic oxaloacetic transaminase (GOT), glutamic pyruvic transaminase (GPT), and lactate dehydrogenase (LDH) were also significantly higher in early-onset PE. The percentage of infants with intrauterine growth restriction was also significantly higher in women with early-onset PE. There were positive correlations between Hsp70 levels and TNF-α, TNFRI, IL-1ß, IL-12, GOT, GPT, LDH, and uric acid concentrations in early-onset PE group. Thus, early-onset PE was associated with greater maternal and fetal impairment. There are differences in pathophysiology between early- and late-onset PE, highlighting by the difference in Hsp70 levels.

Silibinin modulates the NF-κb pathway and pro-inflammatory cytokine production by mononuclear cells from preeclamptic women.

Preeclampsia (PE) is a complication of human pregnancy associated with an intense inflammatory response involving leukocyte activation, as well as elevated production of pro-inflammatory cytokines. The nuclear transcription factor-kappa B (NF-κB) is present in cells of the immune system and is responsible for transcription of genes coding for pro-inflammatory proteins. Silibinin is the main component of silymarin, a polyphenolic extract obtained from fruits and seeds of Silybum marianum with potent hepatoprotective and anti-inflammatory activities. In this study, we assessed whether silibinin modulated NF-κB activity and the production of pro-inflammatory cytokines by peripheral blood mononuclear cells (PBMCs) from preeclamptic patients. PBMC from women with PE, normotensive (NT) pregnant women, and nonpregnant (NP) women were cultured with or without silibinin (5 µM and 50 µM) and 1 µg/mL lipopolysaccharide (LPS) for 18 h. The supernatants were assayed for tumor necrosis factor-alpha (TNF-α) and interleukin-1 beta (IL-1ß) by ELISA. Cells were cultured for 30 min to evaluate NF-κB activity. There was increased endogenous activation of NF-κB as well as TNF-α and IL-1ß release by PBMC in the PE group compared with the NT and NP groups. A positive correlation between NF-κB activity and cytokine production was also observed in the PE group. Silibinin was capable of reducing, at least in part, the levels of NF-κB and cytokines TNF-α and IL-1ß in preeclamptic women. We conclude that silibinin exhibits potent anti-inflammatory activity on PBMC from preeclamptic women by downmodulation of NF-κB activation and inflammatory cytokine production.

Harmonia axyridis: a threat to Brazilian Coccinellidae? / Harmonia axyridis: uma ameaça aos Coccinellidae brasileiros?

Harmonia axyridis (Pallas) (Coleoptera, Coccinellidae) is a polyphagous Asian species, well-known as a classical biological control agent of aphids around the world, introduced probably accidentally in Brazil, sampled for the first time in 2002. It is an important intraguild predator, competing for food with native coccinellids. It was studied H. axyridis alimentary sources and host plants, its abundance compared with native and established species, the influence of abiotic factors and the seasons over the abundance of H. axyridis throughout one year, and discussed the mechanisms which influence the displacement of species. Harmonia axyridis was found in 38 plant species, among them 20 were new records, feeding on 20 aphid species, eight of them new alimentary records. Between 2006/2007, eight Coccinellidae species were collected and H. axyridis was the most abundant (91.23 percent). Harmonia axyridis peak of abundance occurred in August and September 2007, probably influenced by the temperature and food availability. From 1999 to 2007 a reduction and variation in the diversity of collected species of Coccinellidae were observed with the predominance of H. axyridis, which may indicate their displacement.

Harmonia axyridis (Pallas) (Coleoptera, Coccinellidae) é uma espécie asiática, polífaga e reconhecida como agente de controle biológico de afídeos pelo mundo, provavelmente introduzida acidentalmente no Brasil, coletada pela primeira vez em 2002. É um importante predador intraguilda, pois compete por alimento com as espécies nativas de coccinelídeos. Foram estudadas as fontes alimentares de H. axyridis e suas plantas hospedeiras, sua abundância em relação às espécies nativas e estabelecidas, a influência dos fatores abióticos e as estações do ano sobre sua abundância durante um ano e discutidos os mecanismos que influenciam no deslocamento das espécies nativas de Coccinellidae. Além disso, foi testada a influência dos fatores abióticos e as estações do ano sobre sua abundância durante um ano. Harmonia axyridis foi encontrada em 38 espécies de plantas, sendo 20 delas novos registros, alimentando-se de 20 espécies de afídeos, oito novos registros alimentares. Entre os anos de 2006/2007, oito espécies de Coccinellidae foram coletadas e H. axyridis foi a mais abundante (91,23 por cento). O pico de abundância de H. axyridis ocorreu em agosto e setembro de 2007, provavelmente influenciado pela temperatura e disponibilidade de alimento. Entre 1999 e 2007, foram observadas a redução e variação na diversidade de espécies de Coccinellidae coletadas, com a predominância de H. axyridis, o que pode indicar o desalojamento destas espécies.