Environmental and genetic factors may modify or contribute to the phenotypic differences observed in multigenic and monogenic diseases, such as cystic fibrosis (CF). An analysis of modifier genes can be helpful for estimating patient prognosis and directing preventive care. The aim of this study is to determine the association between seven genetic variants of four modifier genes and CF by comparing their corresponding allelic and genotypic frequencies in CF patients (n = 81) and control subjects (n = 104). Genetic variants of MBL2 exon 1 (A, B, C and D), the IL-8 promoter (−251 A/T), the TNFα promoter (TNF1/TNF2), and SERPINA1 (PI*Z and PI*S) were tested in CF patients and control subjects from northeastern Mexico by PCR-RFLP.
The TNF2 allele (P = 0.012, OR 3.43, 95% CI 1.25–9.38) was significantly associated with CF under the dominant and additive models but was not associated with CF under the recessive model. This association remained statistically significant after adjusting for multiple tests using the Bonferroni correction (P = 0.0482). The other tested variants and genotypes did not show any association with the disease.
Citation: Sanchez-Dominguez CN, Reyes-Lopez MA, Bustamante A, Cerda-Flores RM, Villalobos-Torres MdC, Gallardo-Blanco HL, et al. (2014) The Tumor Necrosis Factor α (-308 A/G) Polymorphism Is Associated with Cystic Fibrosis in Mexican Patients. PLoS ONE 9(3): e90945. https://doi.org/10.1371/journal.pone.0090945
Editor: Amit Gaggar, University of Alabama-Birmingham, United States of America
Received: July 15, 2013; Accepted: February 5, 2014; Published: March 6, 2014
Copyright: © 2014 Sanchez-Dominguez et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Funding: This work was supported by grants from the Consejo Nacional de Ciencia y Tecnología CONACyT (62291 and 48497) and UANL’s PAICyT (1648-07) for ROL, Fondo Mixto de Fomento a la Investigación Científica y Tecnológica CONACYT – Gobierno del Estado de Tamaulipas (73578) and SIP-IPN (20080682) for MARL. The authors gratefully acknowledge scholarships from CONACyT, PIFI-IPN and Universia Santander for CNSD. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing interests: The authors have declared that no competing interests exist.
Gene-environment and gene-gene interactions play a role in the phenotypic expression of genetic diseases in individuals harboring the same genotype . Cystic fibrosis (CF) has an estimated incidence of one in 3000 in the Caucasian population, although its frequency may vary in specific subgroups. A newborn screening study conducted in Mexico City revealed two CF-affected newborns among 7193 screened (1∶3597) participants, suggesting a high frequency of CF among Mexicans . Approximately 1900 mutations and variants have been reported in the CF transmembrane conductance regulator (CFTR) gene, with ΔF508 being the most prevalent mutation (50%–60%, http://www.genet.sickkids.on.ca/app). CF primarily involves epithelial cells in the respiratory tract, intestine, pancreas, bladder, and sweat glands; respiratory failure, however, is the major cause of death in CF patients .
Variants in genes that are involved in the inflammatory response have been studied in CF patients based on their potential effects on inflammation and host defense mechanisms. The mannose binding lectin (MBL2) gene encodes a serum acute-phase protein secreted by the liver, resembling the complement component C1q, that leads to opsonization and activation of the complement system through the classical pathway . The serum concentration and complement-triggering activity of MBL depend on single-base mutations in the MBL2 gene –. These mutations may increase the susceptibility of carriers to colonization by bacterial and viral pathogens . The best known genetic variants in exon 1 of the MBL2 gene are Gly54Asp (the B allele, rs1800450), Gly57Glu (the C allele, rs1800451) and Arg52Cys (the D allele, rs5030737), which are together referred to as the O allele. The interleukin 8 (IL-8) gene codes for a member of the CXC chemokine family and is mainly involved in the initiation and amplification of acute inflammatory reactions . IL-8 is produced by a wide range of cell types, such as monocytes, macrophages, and fibroblasts; it primarily mediates the activation and migration of neutrophils from peripheral blood into pathogen-infected tissue, initiating and amplifying inflammatory processes . A polymorphism in position −251 of the IL-8 gene (rs4073) is associated with increased IL-8 expression –. The tumor necrosis factor alpha (TNFα) gene expresses a multifunctional pro-inflammatory cytokine secreted in response to numerous specific stimuli, such as lipopolysaccharides. This molecule induces the release of cytokines IL-6 and IL-8 and increases airway mucus production –. The −308 A TNFα promoter polymorphism (TNF2, rs1800629) has been associated with increased TNFα transcription activity relative to the normal TNF1 allele (−308 G) –. The Alpha-1-antitrypsin (AAT, SERPINA1) gene codes for an acute-phase serine protease glycoprotein that limits tissue self-damage during the inflammatory immune response. AAT deficiency, caused by the S (p.E264V, rs17580) and Z (p.E342K, rs28929474) alleles in the SERPINA1 gene, may induce liver and pulmonary disease . Severe AAT deficiency is a co-dominant autosomal hereditary disorder that clinically resembles early onset pulmonary emphysema, particularly in smokers .
In our Cystic fibrosis clinic, we have a broad range of severity of the CF disease, even with patients carrying the same genotype. No reports on variations in modifier genes in Mexican CF patients have been previously published. Our goal was to explore polymorphisms in genes related to host defense in healthy controls and CF patients from northeastern Mexico to find differences in allelic distribution between both groups. In this study, we report the genotype and allele frequencies of seven genetic variants in four previously reported CF modifier genes: MBL, IL-8, TNFα and AAT.
Materials and Methods
The study was approved by the Research and Ethics Committee of the Universidad Autonoma de Nuevo Leon University Hospital (Registry number BI09-003). After signing written informed consent, blood samples were drawn from 81 CF patients attending the Chronic Lung Disease Prevention and Rehabilitation Center (CEPREP, in Spanish) and from control subjects recruited from the University Hospital and School of Medicine (Universidad Autonoma de Nuevo Leon). Also we collected blood samples from 104 control subjects that met the following inclusion criteria: they agreed to informed consent, they were born in northeastern Mexico (the states of Nuevo Leon, Tamaulipas, Coahuila, and San Luis Potosi), and they belonged to a family with at least three ascending Mexican generations. Genomic DNA was isolated from peripheral venous blood using the phenol–chloroform method, precipitated in ethanol, and finally suspended in Tris-EDTA (pH 7.8).
Screening for the CFTR Gene Mutations
Mutation screening was performed according to the availability of resources and kits along the time. For previously screened CF patients: direct detection of ΔF508 mutation, and Roche ASO16 and 27 mutations kits, for new CF patients: direct detection of ΔF508 mutation, INNOLiPA CFTR36 probe kit and Exon 11-specific PCR and sequencing. Short descriptions of the methodologies are presented below.
PCR and electrophoresis to detect the ΔF508 mutation: PCR product was analyzed in polyachrylamide gels and the diagnosis was established comparing to molecular marker and DNAs of previously ΔF508 diagnosed patients and control subjects (98 bp band for normal allele or 95 bp band for ΔF508 mutation) .
Roche ASO16 and 27 kits (Roche Molecular Systems, Alameda CA, USA) or the INNOLiPA CFTR36 probe kit (Innogenetics, Ghent, Belgium). Methodology consisted of multiplex PCR reactions with biotinylated primers. After verifying the amplification in an agarose gel of 2%, the products were hybridized to membrane bound probes. A positive result was expressed as the appearance of a purple band. Both kits detected normal and mutated versions to report homozygous or heterozygous status for the CF patients. Complete list of mutations is shown in Table 1.
Exon 11- specific PCR and sequencing. Exon 11 PCR fragment was amplified and sequenced in one patient with absence of hybridization of PCR product on the normal and mutated versions of the G551D probe in the CFTR36 probe kit.
Modifier Genes Analysis
DNA from CF patients and control subjects was tested for the Gly54Asp (B allele, rs1800450), Gly57Glu (C allele, rs1800451), Arg52Cys (D allele, rs5030737), and A (wild type) alleles in exon 1 of the MBL2 gene; the −251 T/A (rs4073) allele of the IL-8 gene; the −308 G/A (TNF1/2, rs1800629) alleles of the TNFα gene; and the PI*S Glu264Val (rs17580), PI*Z Glu342Lys (rs28929474), and PI*M (wild type) alleles of the SERPINA1 gene. PCR-RFLP protocols were adapted from previously reported methods. More details are explained in Table 2 and Figure 1 –.
1A: the 134 bp PCR product from exon 1 of the MBL1 gene was digested with Mwo I, Ban I and Mbo II for detection of polymorphisms in the 52, 54 and 57 codons. 1B: the 816 bp PCR product from promoter region of the IL-8 gene was digested with Mfe I for detection of the −251 polymorphism. 1C: the 142 bp PCR product from promoter region of the TNFα gene was digested with Nco I for detection of the −308 polymorphism (TNF2); the 98 bp PCR product from the SERPINA1 gene was digested with Taq Iα for detection of the S genetic variant; the 144 bp PCR product from the SERPINA1 gene was digested with Taq Iα enzyme for detection of the Z genetic variant. Mw1 is the molecular marker pBs+Msp I, Mw2 is the molecular marker λ+Pst I. PMBL, PIL8, PTNF, PAATS and PAATZ are undigested PCR products. The Z allele was not detected.
The SNP & Variation Suite (SVS) 7 (Golden Helix Inc., Bozeman, MA, USA) software program was used to perform all statistical analyses. The association between the tested genotypes and CF was analyzed by correlation/trend and chi-squared (χ2) tests under three different models (dominant, recessive, and additive) and was confirmed with the Bonferroni correction to detect the false discovery rate. Odds ratios were estimated within 95% confidence intervals. Values of P<0.05 were considered statistically significant. The Hardy–Weinberg Equilibrium (HWE) P-values were assessed using a chi-square test.
Eighty-one CF patients and 104 control subjects were recruited for this study. Genotype frequencies for the CFTR gene are described in Table 3. A complete genotype characterization was achieved in 55.6% (n = 45) of the CF patients; in 39.5% (n = 32) of the CF patients, only one mutation was identified, and in 4.9% (n = 4) of the CF patients, both mutations remained undetected. The most prevalent genotypes were ΔF508/other (46.9%, n = 38) and ΔF508/ΔF508 (35.8%, n = 29). The overall frequency of the ΔF508 allele among CF patients was 59.3%. Ten additional mutations were detected: G542X (4.9%), S549N (3.1%), 2789+5G>A (2.5%), 3849+10 kb (1.9%), G85E, R1162X, I148T, R334W, ΔI507, and L206W (0.6% each one). Mutations in the CFTR gene were not detected in 24.7% of the total CFTR alleles.
The PCR-RFLP patterns of the seven genetic variants in the four modifier genes are shown in the Figure 1. The Z allele, and the homozygote TNF2 and AATS genotypes were not found. The genotype frequencies of modifier genes in CF patients and control subjects are shown in Table 4. The B, C, and D alleles of the MBL2 gene were grouped together and reported as the O allele. The polymorphisms were in Hardy-Weinberg equilibrium. The frequencies of the mutant alleles for CF patients and controls were: the MBL-O allele 0.231 and 0.233, the IL-8 -251T allele 0.576 and 0.569, AATS allele 0.012 and 0.014, and TNF2 allele 0.087 and 0.029, respectively. The TNF2 allele (P = 0.012, Odds Ratio (OR) 3.43, 95% CI 1.25–9.38) was significantly associated with CF patients using the dominant model. This association remains statistically significant after adjusting for multiple testing using the Bonferroni correction (P = 0.0482). The association value and the OR of the TNF2 allele were statistically significant when assessed using the additive model (Dd vs. dd) but not the recessive model (Table 5). The other genetic variants tested did not show any association.
In the present study, the ΔF508 mutation accounted for 59.3% of the mutated CFTR alleles, a frequency that resembles the mutation frequencies of European Mediterranean countries. In this regard, this work may be comparable with modifier genes studies performed in those countries. Two previous CFTR mutation reports in the Mexican population showed ΔF508 frequencies of 40.72% , 45%  and 44.6% . Differences could be explained by the clinical criteria, geographic origin and the analytical methods available at that time. The Spanish federation reported a frequency of 45%. Previous studies in the Hispanic population reported a ΔF508 allele frequency ranging from 29 to 46% –. The frequencies of the G542X, R1162X and R334W alleles reflect the Spanish heritage in the Mexican population, but the S549N and 2789+5 G>A alleles are not among the most frequent in Spain –. In this study we previously detected homozygote S549N and ΔF508/S549N genotypes with the ASO16 kit. A second patient was detected by exon 11 direct sequencing because she presented an abnormal pattern of the INNOLiPA CFTR36 kit with the normal and mutant G551D probes. This kit along with the ASO27 excluded the S549N mutation from the mutation panel, hindering the CFTR molecular diagnosis in our population. CFTR gene sequencing should be performed for new or rare CFTR mutations in Mexican population as S549N, and those mutations obtained from central Mexico patients (P750L, 846delT, 4160insGGGG and 297–1 G>A) , as well as for those mutations that remained undetected. It is necessary to establish an adequate diagnosis strategy based in Mexican genetic profile, considering that available commercial kits are designed mainly for Caucasian mutations profile.
In northeastern Mexico, medical care for CF patients is offered at the Cystic Fibrosis Clinic of the CEPREP (http://www.ceprep.edu.mx). Since 1987, approximately 200 patients have been diagnosed with CF based on clinical and molecular analyses. Once the diagnosis of CF is established, the rate of adherence to medical treatment and long-term medical monitoring is low, making investigations into the genetic and environmental factors that influence the outcome in CF Mexican patients difficult.
In this study, we analyzed seven variants in four modifier genes previously reported in CF patients and we found an association between CF and the TNF2 allele.
The proinflammatory role of the TNF2 allele has been demonstrated in B cell line cultures, where the TNF2 allele was more potent transcriptional activator compared to the normal TNF1 . TNF2 allele has been implicated as a potent immunomediator and pro-inflammatory cytokine in the pathogenesis of several human diseases, including pulmonary diseases as CF and asthma. Patients with genotypes related to higher TNFα production had increased frequency of asthma . In Mexican population TNF2 allele was found in 6.0% of asthmatic compared to 2.9% of the controls . These results are similar that those we found in our study (2.9% for controls and 8.9% for CF patients). By the other hand, recent studies in Mexican population reported a higher frequency of the TNF2 allele in healthy unrelated Mexican individuals (7.3%) . Another report related TNF2 allele to breast cancer in Mexican patients compared to healthy women (7.5 and 24.5% respectively) . Differences in TNF1 allele frequency in control subjects could be explained by sample size, methodology and characteristics of the studied group. Our study included population from Northeastern Mexico, while the other studies had different criteria as gender, living in Mexico City or been born in Mexico.
Previous studies in CF patients have shown that TNF2 is associated with a lower percentage of predicted forced expiratory volume in one second (FEV1) and weight z scores . In Mexican population, the TNF2 allele has been associated with rheumatoid arthritis , geriatric lipid profile  and spondyloarthritis . This variant has also been reported to be associated with obesity and asthma –. The high frequency of the TNF2 allele in Mexican CF patients could suggest a heterozygote advantage. In Colombia, an inverse association between the TNF polymorphism and autoimmunity and TB has been reported; this association suggests the existence of a heterozygote advantage and is consistent with the hypothesis that autoimmune diseases are a consequence of natural selection for enhanced TB resistance –.
MBL2, IL-8 and AAT did not show an association with the CF genotype. MBL2 had previously shown associations with different disturbances in the lung function, infection risk, and survival of CF patients , –. In asthma, MBL has been associated with Chlamydia pneumoniae–specific IgG and a greater risk of developing asthma, especially in children with chronic or recurrent infection . MBL levels in asthmatic children positively correlate with peripheral blood eosinophils . MBL therapy may be useful in MBL-deficient patients; it may reduce the susceptibility to or enhance the recovery from bacterial infection or modify the natural history of the disease –. The IL-8 −251 polymorphism has been associated with CF lung disease severity and the differential expression of IL-8, suggesting that the IL-8 variant modifies CF lung disease severity . The −251 variant has been associated with asthma, infection by respiratory syncytial virus, and chronic obstructive pulmonary disease (COPD) , –. Finally, despite the association between AAT deficiency and COPD, studies of AAT variants and infection in CF patients have been inconclusive –. The incidence of AAT deficiency for all five phenotypic classes of the Pi*S and Pi*Z deficiency alleles is 1 in 9.8 for Canada and 1 in 11.3 for the United States. However, a previous report from our group showed very low allele frequencies of Pi*S and Pi*Z variants in a Mexican population (1.5% and 0%, respectively) .
In summary, the frequencies of genetic variants in the MBL2, IL-8, and AAT genes of CF patients did not show significant differences when compared to control subjects, but the TNF2 allele was significantly associated with CF patients. More studies are needed to identify the role of inflammatory mediators in the pathophysiology of CF, as emphasized in previous studies. This information is relevant because clinical trials of drugs targeting TNFα activity  have shown outstanding efficacy in treating chronic inflammatory diseases.
We especially acknowledge the volunteers, whose cooperation made these studies possible. The authors gratefully acknowledge Dr. Sergio Lozano for his critical reading of the manuscript and his contribution in editing the manuscript.
Conceived and designed the experiments: CNSD AB ROL MARL. Performed the experiments: CNSD MCVT. Analyzed the data: HLGB RMCF. Contributed reagents/materials/analysis tools: ROL MARL HLGB. Wrote the paper: CNSD. Helped to design and revised the manuscript: HGMR HABS ARM.
- 1. Collaco JM, Cutting GR (2008) Update on gene modifiers in cystic fibrosis. Curr Opin Pulm Med 14: 559–566.
- 2. Velázquez A, Vela-Amieva M, Naylor EW, Chace DH (2000) Tamiz neonatal ampliado. Rev Mex Pediatr 67: 206–213.
- 3. Rowntree RK, Harris A (2003) The phenotypic consequences of CFTR mutations. Ann Hum Genet 67: 471–485.
- 4. Turner MW (1996) Mannose-binding lectin: the pluripotent molecule of the innate immune system. Immunol Today 17: 532–540.
- 5. Madsen HO, Satz ML, Hogh B, Svejgaard A, Garred P (1998) Different molecular events result in low protein levels of mannan-binding lectin in populations from southeast Africa and South America. J Immunol 161: 3169–3175.
- 6. Yarden J, Radojkovic D, De Boeck K, Macek M Jr, Zemkova D, et al. (2004) Polymorphisms in the mannose binding lectin gene affect the cystic fibrosis pulmonary phenotype. J Med Genet 41: 629–633.
- 7. Garred P, Larsen F, Seyfarth J, Fujita R, Madsen HO (2006) Mannose-binding lectin and its genetic variants. Genes Immun 7: 85–94.
- 8. Eisen DP (2010) Mannose-binding lectin deficiency and respiratory tract infection. J Innate Immun 2: 114–122.
- 9. Harada A, Sekido N, Akahoshi T, Wada T, Mukaida N, et al. (1994) Essential involvement of interleukin-8 (IL-8) in acute inflammation. J Leukoc Biol 56: 559–564.
- 10. Puthothu B, Krueger M, Heinze J, Forster J, Heinzmann A (2006) Impact of IL8 and IL8-receptor alpha polymorphisms on the genetics of bronchial asthma and severe RSV infections. Clin Mol Allergy 4: 2.
- 11. Hull J, Thomson A, Kwiatkowski D (2000) Association of respiratory syncytial virus bronchiolitis with the interleukin 8 gene region in UK families. Thorax 55: 1023–1027.
- 12. Mizunoe S, Shuto T, Suzuki S, Matsumoto C, Watanabe K, et al. (2012) Synergism between interleukin (IL)-17 and toll-like receptor 2 and 4 signals to induce IL-8 expression in cystic fibrosis airway epithelial cells. J Pharmacol Sci 118: 512–520.
- 13. Cowan MJ, Huang X, Yao XL, Shelhamer JH (2000) Tumor necrosis factor alpha stimulation of human Clara cell secretory protein production by human airway epithelial cells. Ann N Y Acad Sci 923: 193–201.
- 14. Lora JM, Zhang DM, Liao SM, Burwell T, King AM, et al. (2005) Tumor necrosis factor-alpha triggers mucus production in airway epithelium through an IkappaB kinase beta-dependent mechanism. J Biol Chem 280: 36510–36517.
- 15. Hajeer AH, Hutchinson IV (2000) TNF-alpha gene polymorphism: clinical and biological implications. Microsc Res Tech 50: 216–228.
- 16. Gambari R, Borgatti M, Lampronti I, Fabbri E, Brognara E, et al. (2012) Corilagin is a potent inhibitor of NF-kappaB activity and downregulates TNF-alpha induced expression of IL-8 gene in cystic fibrosis IB3–1 cells. Int Immunopharmacol 13: 308–315.
- 17. Wilson AG, Symons JA, McDowell TL, McDevitt HO, Duff GW (1997) Effects of a polymorphism in the human tumor necrosis factor alpha promoter on transcriptional activation. Proc Natl Acad Sci U S A 94: 3195–3199.
- 18. Kok KF, te Morsche RH, van Oijen MG, Drenth JP (2010) Prevalence of genetic polymorphisms in the promoter region of the alpha-1 antitrypsin (SERPINA1) gene in chronic liver disease: a case control study. BMC Gastroenterol 10: 22.
- 19. Miravitlles M, Vila S, Jardi R, de la Roza C, Rodriguez-Frias F, et al. (2003) Emphysema due to alpha-antitrypsin deficiency: familial study of the YBARCELONA variant. Chest 124: 404–406.
- 20. Araujo FG, Novaes FC, Santos NP, Martins VC, Souza SM, et al. (2005) Prevalence of deltaF508, G551D, G542X, and R553X mutations among cystic fibrosis patients in the North of Brazil. Braz J Med Biol Res 38: 11–15.
- 21. Sandford AJ, Chagani T, Spinelli JJ, Pare PD (1999) alpha1-antitrypsin genotypes and the acute-phase response to open heart surgery. Am J Respir Crit Care Med 159: 1624–1628.
- 22. Tin SK, Lee LY, Thumboo J, Koh DR, Fong KY (2005) PCR-RFLP genotyping for exon 1 and promoter region mutations of the human mannose binding lectin (MBL-2) gene. J Immunol Methods 303: 148–151.
- 23. Lee WP, Tai DI, Lan KH, Li AF, Hsu HC, et al. (2005) The −251T allele of the interleukin-8 promoter is associated with increased risk of gastric carcinoma featuring diffuse-type histopathology in Chinese population. Clin Cancer Res 11: 6431–6441.
- 24. Chen YP, Pfab T, Slowinski T, Richter CM, Godes M, et al. (2006) Impact of genetic variation of tumor necrosis factor-alpha on gestational hypertension. Chin Med J (Engl) 119: 719–724.
- 25. Orozco L, Velazquez R, Zielenski J, Tsui LC, Chavez M, et al. (2000) Spectrum of CFTR mutations in Mexican cystic fibrosis patients: identification of five novel mutations (W1098C, 846delT, P750L, 4160insGGGG and 297–1G–>A). Hum Genet 106: 360–365.
- 26. Villalobos-Torres C, Rojas-Martinez A, Villareal-Castellanos E, Cantu JM, Sanchez-Anzaldo FJ, et al. (1997) Analysis of 16 cystic fibrosis mutations in Mexican patients. Am J Med Genet 69: 380–382.
- 27. Chavez-Saldana M, Yokoyama E, Lezana JL, Carnevale A, Macias M, et al. (2010) CFTR allelic heterogeneity in Mexican patients with cystic fibrosis: implications for molecular screening. Rev Invest Clin 62: 546–552.
- 28. Grebe TA, Seltzer WK, DeMarchi J, Silva DK, Doane WW, et al. (1994) Genetic analysis of Hispanic individuals with cystic fibrosis. Am J Hum Genet 54: 443–446.
- 29. Sugarman EA, Rohlfs EM, Silverman LM, Allitto BA (2004) CFTR mutation distribution among U.S. Hispanic and African American individuals: evaluation in cystic fibrosis patient and carrier screening populations. Genet Med 6: 392–399.
- 30. Schrijver I, Ramalingam S, Sankaran R, Swanson S, Dunlop CL, et al. (2005) Diagnostic testing by CFTR gene mutation analysis in a large group of Hispanics: novel mutations and assessment of a population-specific mutation spectrum. J Mol Diagn 7: 289–299.
- 31. Aliño-Pellicer SF, Antelo-Landeira MC, Baamonde-Vidarte A, Beltrán-Bengoechea B, Berná-Torres N, et al. (2003) Federación Española contra la Fibrosis Quística. Libro Blanco de atención a la fibrosis quística; Consumo MdSy, editor. Madrid, España.
- 32. Alonso MJ, Heine-Suner D, Calvo M, Rosell J, Gimenez J, et al. (2007) Spectrum of mutations in the CFTR gene in cystic fibrosis patients of Spanish ancestry. Ann Hum Genet 71: 194–201.
- 33. Shmarina G, Pukhalsky A, Petrova N, Zakharova E, Avakian L, et al. (2013) TNF gene polymorphisms in cystic fibrosis patients: contribution to the disease progression. J Transl Med 11: 19.
- 34. Jimenez-Morales S, Velazquez-Cruz R, Ramirez-Bello J, Bonilla-Gonzalez E, Romero-Hidalgo S, et al. (2009) Tumor necrosis factor-alpha is a common genetic risk factor for asthma, juvenile rheumatoid arthritis, and systemic lupus erythematosus in a Mexican pediatric population. Hum Immunol 70: 251–256.
- 35. Vargas-Alarcon G, Ramirez-Bello J, Juarez-Cedillo T, Ramirez-Fuentes S, Carrillo-Sanchez S, et al. (2012) Distribution of the IL-1RN, IL-6, IL-10, INF-gamma, and TNF-alpha Gene Polymorphisms in the Mexican Population. Genet Test Mol Biomarkers 16: 1246–1253.
- 36. Gomez Flores-Ramos L, Escoto-De Dios A, Puebla-Perez AM, Figuera-Villanueva LE, Ramos-Silva A, et al. (2013) Association of the tumor necrosis factor-alpha −308 G>A polymorphism with breast cancer in Mexican women. Genet Mol Res 12: 5680–5693.
- 37. Hull J, Thomson AH (1998) Contribution of genetic factors other than CFTR to disease severity in cystic fibrosis. Thorax 53: 1018–1021.
- 38. Rodriguez-Carreon AA, Zuniga J, Hernandez-Pacheco G, Rodriguez-Perez JM, Perez-Hernandez N, et al. (2005) Tumor necrosis factor-alpha −308 promoter polymorphism contributes independently to HLA alleles in the severity of rheumatoid arthritis in Mexicans. J Autoimmun 24: 63–68.
- 39. Parra-Rojas I, Ruiz-Madrigal B, Martinez-Lopez E, Panduro A (2006) Influence of the −308 TNF-alpha and −174 IL-6 polymorphisms on lipid profile in Mexican subjects. Hereditas 143: 167–172.
- 40. Vargas-Alarcon G, Casasola-Vargas J, Rodriguez-Perez JM, Huerta-Sil G, Perez-Hernandez N, et al. (2006) Tumor necrosis factor-alpha promoter polymorphisms in Mexican patients with spondyloarthritis. Hum Immunol 67: 826–832.
- 41. Castro-Giner F, Kogevinas M, Imboden M, de Cid R, Jarvis D, et al. (2009) Joint effect of obesity and TNFA variability on asthma: two international cohort studies. Eur Respir J 33: 1003–1009.
- 42. Wu H, Romieu I, Sienra-Monge JJ, del Rio-Navarro BE, Anderson DM, et al. (2007) Parental smoking modifies the relation between genetic variation in tumor necrosis factor-alpha (TNF) and childhood asthma. Environ Health Perspect 115: 616–622.
- 43. Correa PA, Gomez LM, Anaya JM (2004) [Polymorphism of TNF-alpha in autoimmunity and tuberculosis]. Biomedica 24 Supp 1: 43–51.
- 44. Correa PA, Gomez LM, Cadena J, Anaya JM (2005) Autoimmunity and tuberculosis. Opposite association with TNF polymorphism. J Rheumatol 32: 219–224.
- 45. Buranawuti K, Boyle MP, Cheng S, Steiner LL, McDougal K, et al. (2007) Variants in mannose-binding lectin and tumour necrosis factor alpha affect survival in cystic fibrosis. J Med Genet 44: 209–214.
- 46. Muhlebach MS, MacDonald SL, Button B, Hubbard JJ, Turner ML, et al. (2006) Association between mannan-binding lectin and impaired lung function in cystic fibrosis may be age-dependent. Clin Exp Immunol 145: 302–307.
- 47. Davies JC, Turner MW, Klein N (2004) Impaired pulmonary status in cystic fibrosis adults with two mutated MBL-2 alleles. Eur Respir J 24: 798–804.
- 48. Haerynck F, Van Steen K, Cattaert T, Loeys B, Van Daele S, et al. (2012) Polymorphisms in the lectin pathway genes as a possible cause of early chronic Pseudomonas aeruginosa colonization in cystic fibrosis patients. Hum Immunol 73: 1175–1183.
- 49. Nagy A, Kozma GT, Keszei M, Treszl A, Falus A, et al. (2003) The development of asthma in children infected with Chlamydia pneumoniae is dependent on the modifying effect of mannose-binding lectin. J Allergy Clin Immunol 112: 729–734.
- 50. Uguz A, Berber Z, Coskun M, Halide Akbas S, Yegin O (2005) Mannose-binding lectin levels in children with asthma. Pediatr Allergy Immunol 16: 231–235.
- 51. Garred P, Pressler T, Lanng S, Madsen HO, Moser C, et al. (2002) Mannose-binding lectin (MBL) therapy in an MBL-deficient patient with severe cystic fibrosis lung disease. Pediatr Pulmonol 33: 201–207.
- 52. Summerfield JA (2003) Clinical potential of mannose-binding lectin-replacement therapy. Biochem Soc Trans 31: 770–773.
- 53. Kilpatrick DC (2003) Introduction to mannan-binding lectin. Biochem Soc Trans 31: 745–747.
- 54. Hillian AD, Londono D, Dunn JM, Goddard KA, Pace RG, et al. (2008) Modulation of cystic fibrosis lung disease by variants in interleukin-8. Genes Immun 9: 501–508.
- 55. Heinzmann A, Ahlert I, Kurz T, Berner R, Deichmann KA (2004) Association study suggests opposite effects of polymorphisms within IL8 on bronchial asthma and respiratory syncytial virus bronchiolitis. J Allergy Clin Immunol 114: 671–676.
- 56. Arinir U, Klein W, Rohde G, Stemmler S, Epplen JT, et al. (2005) Polymorphisms in the interleukin-8 gene in patients with chronic obstructive pulmonary disease. Electrophoresis 26: 2888–2891.
- 57. Mahadeva R, Stewart S, Bilton D, Lomas DA (1998) Alpha-1 antitrypsin deficiency alleles and severe cystic fibrosis lung disease. Thorax 53: 1022–1024.
- 58. Mahadeva R, Westerbeek RC, Perry DJ, Lovegrove JU, Whitehouse DB, et al. (1998) Alpha1-antitrypsin deficiency alleles and the Taq-I G–>A allele in cystic fibrosis lung disease. Eur Respir J 11: 873–879.
- 59. Meyer P, Braun A, Roscher AA (2002) Analysis of the two common alpha-1-antitrypsin deficiency alleles PiMS and PiMZ as modifiers of Pseudomonas aeruginosa susceptibility in cystic fibrosis. Clin Genet 62: 325–327.
- 60. Frangolias DD, Ruan J, Wilcox PJ, Davidson AG, Wong LT, et al. (2003) Alpha 1-antitrypsin deficiency alleles in cystic fibrosis lung disease. Am J Respir Cell Mol Biol 29: 390–396.
- 61. Sanchez-Dominguez CN, Buenfil-Lozano JA, Molina-Guajardo CA, Borjas-Almaguer OD, Castillo-Lartigue A, et al. (2008) Frequency of S and Z alleles for alpha-1-antitrypsin and tumor necrosis factor alpha −308 promoter polymorphism in northeastern Mexico. Allergy Asthma Proc 29: 406–410.
- 62. Kim J, Remick DG (2007) Tumor necrosis factor inhibitors for the treatment of asthma. Curr Allergy Asthma Rep 7: 151–156.