Farnesyl Diphosphate Synthase Gene Associated with Loss of Bone Mass Density and Alendronate Treatment Failure in Patients with Primary Osteoporosis.

Aminobisphosphonates (NBPs) are the first-choice medication for osteoporosis (OP); NBP treatment aims at increasing bone mineral density (BMD) by inhibiting the activity of farnesyl diphosphate synthase (FDPS) enzyme in osteoclasts. Despite its efficacy, inadequate response to the drug and side effects have been reported. The A allele of the rs2297480 (A > C) SNP, found in the regulatory region of the FDPS gene, is associated with reduced gene transcription. This study evaluates the FDPS variant rs2297480 (A > C) association with OP patients' response to alendronate sodium treatment. A total of 304 OP patients and 112 controls were enrolled; patients treated with alendronate sodium for two years were classified, according to BMD variations at specific regions (lumbar spine (L1-L4), femoral neck (FN) and total hip (TH), as responders (OP-R) (n = 20) and non-responders (OP-NR) (n = 40). We observed an association of CC genotype with treatment failure (p = 0.045), followed by a BMD decrease in the regions L1-L4 (CC = -2.21% ± 2.56; p = 0.026) and TH (CC = -2.06% ± 1.84; p = 0.015) after two years of alendronate sodium treatment. Relative expression of the FDPS gene was also evaluated in OP-R and OP-NR patients. Higher expression of the FDPS gene was also observed in OP-NR group (FC = 1.84 ± 0.77; p = 0.006) when compared to OP-R. In conclusion, the influence observed of FDPS expression and the rs2897480 variant on alendronate treatment highlights the importance of a genetic approach to improve the efficacy of treatment for primary osteoporosis.

Vitamin D receptor gene polymorphisms influence on clinical profile and bone mineral density at different skeletal sites in postmenopausal osteoporotic women.

Bone remodeling is marked by bone synthesis and absorption balance, and any altered dynamic in this process leads to osteoporosis (OP). The interaction of hormonal, environmental and genetic factors regulate bone metabolism. Since vitamin D displays a classic role in bone metabolism regulation, acting through vitamin D receptor (VDR), the genetic variants within VDR were the first ones associated with bone density and remodelling. Therefore, we investigated whether three single nucleotide polymorphisms (SNPs) within VDR were associated with OP differential susceptibility and clinical profile from postmenopausal versus healthy women from Northeast Brazil. Genetic association study enrolling 146 postmenopausal osteoporotic women as the patient group and 95 healthy age-matched women as the control group. We assessed three SNPs within VDR (rs11168268, rs1540339 and rs3890733), considering the clinical profile of all patients. Our results showed an association of rs11168268 G/G genotype with higher bone mineral density (BMD) mean for the total hip (A/A = 0.828 ± 0.09; A/G = 0.081 ± 0.13; G/G = 0.876 ± 0.12, p = .039), and the rs3890733 T/T genotype was associated with increased OP risk in patients below 60 years old (odds ratio [OR] = 5.12, 95% confidence interval [CI ]= 1.13-23.27, p = .012). The rs1540339 T/T genotype was associated with protection for individuals with low melanin deposition when compared to the high melanin deposition group (OR = 0.24, 95%CI = 0.06-0.94, p = .029). Additionally, 61% of patients presented deficient vitamin D serum levels. The SNP rs11168268 G/G was associated with a significantly increased mean total hip BMD in patients OP, highlighting this SNP and its relationship with BMD.

Can Polymorphisms in NLRP3 Inflammasome Complex Be Associated with Postmenopausal Osteoporosis Severity?

The immune system plays a critical role in bone homeostasis and, consequently, in the pathophysiology of postmenopausal osteoporosis (OP) since estrogen deficiency induces the inflammasome and increases production of pro-inflammatory cytokines, such as IL-1ß and IL-18. NLRP3 inflammasome complex genes have been related with bone homeostasis in cellular and animal models. Here, we performed an association study evaluating SNVs (single-nucleotide variants) in inflammasome NLRP3 pathway genes (NLRP3, CARD8, CASP1, IL-18, and IL-1ß) to assess whether variants in these genes could be related to susceptibility to primary OP in postmenopausal women. METHODS: We genotyped 196 postmenopausal OP patients and 103 healthy controls using SNV-specific Taqman® probes. Data and statistical analyses were performed using the SNPstats and GraphPad Prism 8 software. RESULTS: We showed an association between NLRP3 rs35829419 CA genotype and lower bone mineral density (BMD) mean at the lumbar spine (p = 0.001); we also observed an association between IL-1ß rs16944 AA genotype and higher BMD mean at the total hip (p = 0.009). The IL-1ß rs16944 GG was associated with lower alkaline phosphatase levels (ALP) (p = 0.009), and the IL-18 rs1946519 AA was associated with lower vitamin D levels (p = 0.018). Additionally, OP patients presented deficient vitamin D and parathyroid hormone (PTH). CONCLUSIONS: The NLRP3 inflammasome complex SNVs were associated with OP severity, possibly indicating these genes' participation in bone metabolism and its dysregulation.

Expression of the miR-9-5p, miR-125b-5p and its target gene NFKB1 and TRAF6 in childhood-onset systemic lupus erythematosus (cSLE).

Childhood- onset systemic lupus erythematosus (cSLE) is a multisystem inflammatory disease that can lead to severe clinical conditions resulting in early comorbidities. Several genetic, environmental, and immunological factors are known to influence the onset of the disease. MiRNAs have been already considered as potential actors involved in the development and activity of the SLE. Thus, understanding the behavior of these regulators can contribute to clarify the inflammatory process affecting SLE patients. Among miRNAs, miR-125b-5p and miR-9-5p targeting NFKB1 and TRAF6 genes can be involved in the etio-pathogenesis of the disease by modulating inflammation. In this study we evaluated miR-9-5p and miR-125b-5p expression and its target genes NFKB1 and TRAF6 in peripheral blood samples (PBMC) from the 35 cSLE patients and 35 healthy controls. MiRNAs and gene target expression have been evaluated by using RT-PCR with specific TaqMan® probes. Both miR-9-5p [Fold Change (FC) = -2.21; p = 0.002] and miR-125b-5p (FC= -3.30; p < 0.0001) and NFKB1 (FC = -1.84; p < 0.001) were downregulated in cSLE patients, while TRAF6 was upregulated (FC = 1.80; p = 0.006) in cSLE patients when compared to controls. A significant correlation was found between miR-125b-5p and its target gene NFKB1 [Spearman (r) = 0.47; p = 0.023]. Our results showed miR-125b-5p and miR-9-5p differential expression in cSLE patients, possibly contributing to better understanding the role of these regulators in cSLE development and disease pathogenesis.

Relação do processamento sensorial e sistema estomatognático de crianças respiradoras orais / Relation of sensory processing and stomatognical system of oral respiratory children

RESUMO Objetivo Verificar a relação entre o processamento sensorial e as alterações das funções do Sistema Estomatognático de crianças respiradoras orais, caracterizando o processamento sensorial destas e comparando-o com o de respiradoras nasais. Método Foram selecionadas 50 crianças (5 a 12 anos) que apresentaram diagnóstico de respiração oral e 50 sem sinais e sintomas de respiração oral ou rinite alérgica para fazer parte do grupo controle, pareadas por idade e sexo. As crianças respiradoras orais e nasais passaram por avaliação do processamento sensorial, através da Sensory Processing Measure - Home form, e as respiradoras orais por avaliação da motricidade orofacial através da Avaliação Miofuncional Orofacial com Escore. Os resultados foram apresentados em forma de tabela e com suas respectivas frequências absoluta e relativa. Resultados A maioria das crianças avaliadas foi do sexo masculino, estando com idade média de 8 anos. A maioria dos respiradores orais apresentou alteração no processamento de todos os sentidos, com relação estatisticamente significativa quando comparados com os respiradores nasais. Houve relação, nos respiradores orais, entre o processamento sensorial proprioceptivo e o movimento das bochechas, processamento sensorial visual e movimentação da cabeça durante a deglutição e entre o tipo de mastigação e o processamento sensorial tátil. Conclusão Após análise dos dados foi possível perceber que o processamento sensorial de todos os sistemas se apresenta com alteração nos respiradores orais e que esse mau processamento se relaciona a mobilidade orofacial, bem como com funções do Sistema Estomatognático, além do tipo de mastigação dessa população.

ABSTRACT Purpose To verify the relationship between sensory processing and changes in the functions of the stomatognathic system in mouth breathing children, characterizing their sensory processing and comparing it with that of nasal breathing children. Methods 50 children (5 to 12 years) who were diagnosed with mouth breathing and 50 without signs and symptoms of mouth breathing or allergic rhinitis were selected to be part of the control group, matched for age and sex. Oral and nasal breathing children underwent sensory processing evaluation, through the Sensory Processing Measure - home form, and mouth breathers, through the evaluation of orofacial motricity through the Orofacial Myofunctional Evaluation with score. The results were presented in table form and with their respective absolute and relative frequencies. Results Most of the children evaluated were male, with an average age of eight years. Most mouth breathers presented alteration in the processing of all senses, with a statistically significant relationship when compared to nasal breathers. There was a relationship, in mouth breathers, between proprioceptive sensory processing and the movement of the cheeks, visual sensory processing and head movement during swallowing, and between the type of chewing and tactile sensory processing. Conclusion After analyzing the data, it was possible to see that the sensory processing of all systems presents with changes in mouth breathers and that this poor processing is related to orofacial mobility, as well as functions of the stomatognathic system, in addition to the type of chewing of this population.

The Role of NLRP3 Inflammasome in Lupus Nephritis.

Lupus nephritis (LN) is the most frequent and severe of systemic lupus erythematosus (SLE) clinical manifestations and contributes to the increase of morbidity and mortality of patients due to chronic kidney disease. The NLRP3 (NLR pyrin domain containing 3) is a member of the NLR (NOD-like receptors), and its activation results in the production of pro-inflammatory cytokines, which can contribute to the pathogenesis of LN. In this review manuscript, we approach the relation between the NLRP3 inflammasome, SLE, and LN, highlighting the influence of genetic susceptibility of NLRP3 polymorphisms in the disease; the main functional studies using cellular and animal models of NLRP3 activation; and finally, some mechanisms of NLRP3 inhibition for the development of possible therapeutic drugs for LN.

Relation of sensory processing and stomatognical system of oral respiratory children. / Relação do processamento sensorial e sistema estomatognático de crianças respiradoras orais.

PURPOSE: To verify the relationship between sensory processing and changes in the functions of the stomatognathic system in mouth breathing children, characterizing their sensory processing and comparing it with that of nasal breathing children. METHODS: 50 children (5 to 12 years) who were diagnosed with mouth breathing and 50 without signs and symptoms of mouth breathing or allergic rhinitis were selected to be part of the control group, matched for age and sex. Oral and nasal breathing children underwent sensory processing evaluation, through the Sensory Processing Measure - home form, and mouth breathers, through the evaluation of orofacial motricity through the Orofacial Myofunctional Evaluation with score. The results were presented in table form and with their respective absolute and relative frequencies. RESULTS: Most of the children evaluated were male, with an average age of eight years. Most mouth breathers presented alteration in the processing of all senses, with a statistically significant relationship when compared to nasal breathers. There was a relationship, in mouth breathers, between proprioceptive sensory processing and the movement of the cheeks, visual sensory processing and head movement during swallowing, and between the type of chewing and tactile sensory processing. CONCLUSION: After analyzing the data, it was possible to see that the sensory processing of all systems presents with changes in mouth breathers and that this poor processing is related to orofacial mobility, as well as functions of the stomatognathic system, in addition to the type of chewing of this population.

OBJETIVO: Verificar a relação entre o processamento sensorial e as alterações das funções do Sistema Estomatognático de crianças respiradoras orais, caracterizando o processamento sensorial destas e comparando-o com o de respiradoras nasais. MÉTODO: Foram selecionadas 50 crianças (5 a 12 anos) que apresentaram diagnóstico de respiração oral e 50 sem sinais e sintomas de respiração oral ou rinite alérgica para fazer parte do grupo controle, pareadas por idade e sexo. As crianças respiradoras orais e nasais passaram por avaliação do processamento sensorial, através da Sensory Processing Measure ­ Home form, e as respiradoras orais por avaliação da motricidade orofacial através da Avaliação Miofuncional Orofacial com Escore. Os resultados foram apresentados em forma de tabela e com suas respectivas frequências absoluta e relativa. RESULTADOS: A maioria das crianças avaliadas foi do sexo masculino, estando com idade média de 8 anos. A maioria dos respiradores orais apresentou alteração no processamento de todos os sentidos, com relação estatisticamente significativa quando comparados com os respiradores nasais. Houve relação, nos respiradores orais, entre o processamento sensorial proprioceptivo e o movimento das bochechas, processamento sensorial visual e movimentação da cabeça durante a deglutição e entre o tipo de mastigação e o processamento sensorial tátil. CONCLUSÃO: Após análise dos dados foi possível perceber que o processamento sensorial de todos os sistemas se apresenta com alteração nos respiradores orais e que esse mau processamento se relaciona a mobilidade orofacial, bem como com funções do Sistema Estomatognático, além do tipo de mastigação dessa população.

Genetic variants in LGALS3 are related to lower galectin-3 serum levels and clinical outcomes in systemic sclerosis patients: A case-control study.

INTRODUCTION: Systemic sclerosis (SSc) is a rare complex disease characterized by vascular damage, autoimmunity, and extensive skin and internal organs fibrosis. Galectin-3 (Gal-3) is encoded by gene LGALS3 (Lectin, Galactoside-Binding, Soluble, 3; 14q22.3) and it has been reported to play a central role in self-tolerance, inflammation, and fibrosis. OBJECTIVE: To investigate associations among LGALS3 single nucleotide polymorphisms (SNPs) and serum levels Gal-3 and SSc susceptibility and their clinical features. METHODS: A case-control study with 88 patients and 151 matched controls was performed. LGALS3 variants were analyzed by the TaqMan real-time polymerase chain reaction (PCR) system whereas Gal-3 serum levels were measured by sandwich enzyme linked immunosorbent assay (ELISA). Associations among genotypes, clinical features, and Gal-3 levels were performed by univariable and multivariable analysis through statistical packages. RESULTS: The LGALS3 rs4652 A/C genotype was more frequent in SSc patients than controls according to overdominant model [OR 1.89 (CI 95% 1.01 - 3.52); p = .046]. Also, LGALS3 rs4652 C/C polymorphic genotype was associated with lower patient Gal-3 levels (p = .03) and control group (p = 0.005), as noted by generalized linear model (GLM). The LGALS3 rs1009977 G/T controls showed higher Gal-3 levels than wild-type and polymorphic genotypes (p = .03); however, in SSc patients, no difference was found. None of the LGALS3 SNPs or Gal-3 levels was associated with clinical manifestations in SSc patients. Considering only the SSc group, GLM analysis pointed LGALS3 rs4652 and rs2075601, pulmonary arterial hypertension (PAH), myopathy, and health assessment questionnaire (HAQ) and scleroderma health assessment questionnaire (SHAQ) as important predictors for Gal-3 levels. CONCLUSION: The LGALS3 rs4652 A/C was more frequent in SSc patients and related to lower Gal-3 levels. These findings were corroborated through a GLM to estimate Gal-3 values. Also, by model equations, Gal-3 levels may be predicted by HAQ, SHAQ, PAH, myopathy, and LGALS3 rs4652 and rs2075601 factors. In these ways, we suggest that galectins may be promising biomarkers to identify susceptibility to SSc as well as to identify HAQ, SHAQ, PAH, and myopathy outcomes.

Is there an Inflammation Role for MYD88 in Rheumatoid Arthritis?

Rheumatoid arthritis (RA) is an autoimmune and inflammatory disease with strong genetic influence, especially upon immune response components. Several cytokines from the toll-like receptors activation pathway display recognized role for RA establishment. However, few studies have verified the role of key mediators such as MYD88 gene and its genetic variants. In the present study, we aim to evaluate the rs6853 functional single-nucleotide variation (SNV) role in RA etiopathogenesis, clinical severity status, and its impact in MYD88 mRNA levels and IL-lß protein levels. For the association study, a total of 423 RA patients and 346 health individuals, enrolled as control, from Northeast and Southeast Brazil were genotyped using specific Taqman probe. For the gene expression assays, we performed a MYD88 rs6853 genotype-guided monocyte cell culture divided into non-stimulated and lypopolysaccharides (LPS)-stimulated cells from healthy individuals. MYD88 gene expression was measured using primer specifics while IL-1ß levels were evaluated by ELISA. We observed that A allele and AA genotype were associated to an increased risk to RA development (OR = 1.60; 95% CI 1.24-2.08; p = 0.0004/OR = 2.83; 95% CI 1.25-6.41; p = 0.0152). The AA genotype exhibited lower MYD88 mRNA levels than GG genotype in non-stimulated monocyte cell culture (FC - 3.83; p = 0.003). Additionally, we verified an increase of IL-1ß levels when AA genotype non-stimulated monocytes were compared to AA genotype LPS-stimulates (p = 0.021). In summary, MYD88 rs6853 polymorphism associated to RA development in our Brazilian cohort and showed influence upon MYD88 mRNA levels' expression and IL-lß production.

High burden of acute kidney injury in COVID-19 pandemic: systematic review and meta-analysis.

AIMS: Hospitalised patients with COVID-19 have a variable incidence of acute kidney injury (AKI) according to studies from different nationalities. The present systematic review and meta-analysis describes the incidence of AKI, need for renal replacement therapy (RRT) and mortality among patients with COVID-19-associated AKI. METHODS: We systematically searched electronic database PubMed, SCOPUS and Web of Science to identify English articles published until 25 May 2020. In case of significant heterogeneity, the meta-analyses were conducted assuming a random-effects model. RESULTS: From 746 screened publications, we selected 21 observational studies with 15 536 patients with COVID-19 for random-effects model meta-analyses. The overall incidence of AKI was 12.3% (95% CI 7.3% to 20.0%) and 77% of patients with AKI were critically ill (95% CI 58.9% to 89.0%). The mortality among patients with AKI was 67% (95% CI 39.8% to 86.2%) and the risk of death was 13 times higher compared with patients without AKI (OR=13.3; 95% CI 6.1 to 29.2). Patients with COVID-19-associated AKI needed for RRT in 23.4% of cases (95% CI 12.6% to 39.4%) and those cases had high mortality (89%-100%). CONCLUSION: The present study evidenced an incidence of COVID-19-associated AKI higher than previous meta-analysis. The majority of patients affected by AKI were critically ill and mortality rate among AKI cases was high. Thus, it is extremely important for health systems to be aware about the impact of AKI on patients' outcomes in order to establish proper screening, prevention of additional damage to the kidneys and adequate renal support when needed.

Differential expression of the inflammasome complex genes in systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is a complex autoimmune disorder involving heterogeneous clinical manifestations and numerous susceptibility genes. Several findings evidence the critical role of inflammasomes in the predisposition to autoimmune diseases and in SLE. We investigated whether inflammasome polymorphins could affect susceptibility to develop and/or severity SLE. Moreover, differences in inflammasome activation in peripheral blood were also evaluated in SLE patients and controls. The distribution of 13 SNPs in eight inflammasome genes was evaluated. To assess inflammasome priming in peripheral blood monocytes of SLE and controls, differential expression of selected inflammasome genes and IL-1ß production was analyzed in resting condition as well as after LPS and ATP stimulation. Results showed that the gain-of-function variant rs10754558 (NLRP3) was significantly more frequent in SLE patients with nephritis, reinforcing the concept of a key role of NLRP3 inflammasome not only in SLE but also especially in kidney disease. SLE monocytes in resting condition showed a higher level of IL-1ß expression and produced higher levels of IL-1ß when stimulated with LPS+ATP comparing to controls. The stimulation induced a significant expression of NLRP1, AIM2, CASP1, and IL1B genes, suggesting that the NLRP1 inflammasome is responsible for the IL-1ß production observed in monocytes. These data emphasized once more the important contribution of inflammasome in SLE-associated inflammation.

IL1ß, IL18, NFKB1 and IFNG gene interactions are associated with severity of rheumatoid arthritis: A pilot study.

Rheumatoid arthritis (RA) is an autoimmune disease which can lead to progressive and functional disability. Literature data suggest that some inflammatory proteins are dysregulated in RA patients and its genetic polymorphisms may contribute to the aetiology and pathogenesis of disease in different ethnic groups. Polymorphisms in IL1ß, IL18, NFKB1 and IFNG genes were studied in different populations with RA, but the analysis indicated contradictory results. Thereby, we hypothesised that polymorphisms in these genes could have a combined effect on susceptibility to and severity of disease. We evaluated the +3953 C/T IL1ß (rs1143634), -137 G/C IL18 (rs187238), -94 ins/del ATTG NFKB1 (rs28362491) and +874 T/A IFNG (rs2430561) polymorphisms in the northeastern Brazilian population. Peripheral blood samples were collected and DNA extraction was conducted. The polymorphisms were evaluated by RFLP and ARMS-PCR. An association was observed in rs1143634 which showed a protective effect against development of RA in carriers of the T allele (OR = 0.58; 95% CI 0.36-0.92; p = .020). In addition, we found an association among genotypes of the rs1143634 with the HAQ index (p = .021) and rs2430561 with DAS28 (p = .029) and CDAI (p = .029). In relation to combined effects of these SNPs (C/C to rs1143634, G/G to rs187238, I/I to rs28362491 and AA to rs2430561) we found a significant association with decreased functional disability (HAQ index p < .001) and ESR (p = .034), indicating a lower disease activity in carriers of these genotypes. GLM analysis confirmed these associations (HAQ (F = 5.497; p < .001) and ESR (F = 2.727; p = .032)). Our analysis indicated that in the studied population +3953 C/T IL-1ß (rs1143634), -137 G/C IL-18 (rs187238), -94 ins/del ATTG NFKB1 (rs28362491) and +874 T/A IFNG (rs2430561) polymorphisms can together contribute to RA severity although they do not individually influence the disease.

Are key cytokines genetic and serum levels variations related to rheumatoid arthritis clinical severity?

This study evaluated the possible association between SNPs in cytokines coding genes, namely IL10, IL6 and IFNG, cytokines serum levels and clinical assessment' scores in patients with Rheumatoid Arthritis(RA). SNPs genotyping was performed in 126 RA patients and 177 healthy individuals with Taqman probes specific for IL10 -1082 (T>C, rs1800896);INFG -1616 (A>G, rs2069705) and IL6 -174 (G>C, rs1800795) variants,positioned in regulatory regions. Cytokine Bead Array (CBA) was used to measure cytokine levels. We found association between INFG -1616 G allele(p = 0.0210; OR = 1.605) and INFG -1616 GG genotype (p = 0.0268; OR =2.609) and RA susceptibility. We also observed association between IL10 -1082 TT genotype and high clinical disease activity index (CDAI) values (p = 0.026; OR = 1.906; 95% CI = 1.082 - 3.359), IL10 -1082 CC genotype and low CDAI values (p = 0.016; OR = 0.256) and INFG -1616 AA and high CDAI values (p = 0.025; OR = 2.919). IL10 -1082 CC also exhibited the lowest IL-10 levels than IL10 -1082 TT (p = 0.020) and IL10 -1082 TC (p = 0.032). Finally, we verified higher IL-6 value in the RA patients than healthy control group (p = 0.007) and an association between high IL-6 levels and increased CDAI (r = 0.4648, p = 0.0015); DAS 28 (r = 0.3933, p= 0.0091), presence of bone erosions (r = 0.3170, p = 0.0361), ESR levels(r = 0.3041, p = 0.0448) and IFN-γ levels (r = 0.3049, p = 0.0468).Altogether, we suggest that IL10 -1082 (T>C, rs1800896) and INFG -1616(A>G, rs2069705) polymorphisms as well as IL-6 levels alterations may play a role for prognostic and disease follow-up.

Postmenopausal Osteoporosis reference genes for qPCR expression assays.

Osteoporosis (OP) is a multifactorial disease influenced by genetic factors in more than half of the cases. In spite of the efforts to clarify the relationship among genetic factors and susceptibility to develop OP, many genetic associations need to be further functionally validated. Besides, some limitations as the choice of stably expressed reference genes (RG) should be overcome to ensure the quality and reproducibility of gene expression assays. To our knowledge, a validation study for RG in OP is still missing. We compared the expression levels, using polymerase chain reaction quantitative real time (qPCR) of 10 RG (G6PD, B2M, GUSB, HSP90, EF1A, RPLP0, GAPDH, ACTB, 18 S and HPRT1) to assess their suitability in OP analysis by using GeNorm, Normfinder, BestKeeper and RefFinder programs. A minimal number of two RG was recommended by GeNorm to obtain a reliable normalization. RPLP0 and B2M were identified as the most stable genes in OP studies while ACTB, 18 S and HPRT1 were inadequate for normalization in our data set. Moreover, we showed the dramatic effects of suboptimal RG choice on the quantification of a target gene, highlighting the importance in the identification of the most appropriate reference gene to specific diseases. We suggest the use of RPLP0 and B2M as the most stable reference genes while we do not recommend the use of the least stable reference genes HPRT1, 18 S and ACTB in OP expression assays using PBMC as biological source. Additionally, we emphasize the importance of individualized and careful choice in software and reference genes selection.

IFN-gamma and IL-12B polymorphisms in women with cervical intraepithellial neoplasia caused by human papillomavirus.

The Human Papillomavirus (HPV) is a sexually transmitted organism associated with Cervical Intraepithelial Neoplasia (CIN) and cervical cancer, the second main cause of malignancy in women worldwide. The virus itself, however, is not enough to cause lesions on the cervix. Several studies suggest that some polymorphic sites changes the cytokines levels and influence the cancer development in HPV infected patients. In this study, we evaluated the presence of functional polymorphisms at +874 (T/A) IFNG and +1188 (A/C) IL-12B genes in cervical smears samples from 76 healthy women and 162 women, HPV positive, with CIN lesion--CIN I (45), CIN II (55), CIN III (53) and cervical cancer (9)--in Brazilian population. There was no significant differences in genotype (p = 0.4192) and allele (p = 0.370; OR = 1.20) distributions between CIN patients and control groups on IFNG allelic polymorphism. Moreover, for IL-12B gene, there was a significant difference in genotype (p = 0.015) and allele distribution (p = 0.014; OR = 0.5754) between the groups. When samples were stratified according to grade of cervical lesion, the AA genotype and A allele were less frequent in the group with low-grade cervical lesions than in group with high-grade cervical lesions (p = 0.0036 and p = 0.0010; OR = 0.3819, respectively), suggesting that the C allele (mutant) may protect against the emergence of CIN lesions and its progression.