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PURPOSE: Targeted axillary dissection (TAD) after neoadjuvant therapy (NAT) includes removing of marked and sentinel lymph nodes (SLNs). The aim was to investigate the optimization of TAD localization techniques after NAT among breast cancer patients. METHODS: From November 2020 to 2022, we prospectively enrolled 107 lymph node-positive breast cancer patients in XX Hospital and received complete cycles of NAT. Patients were randomly divided into the following 3 groups before treatment: group A, marked node with clip (n=34); group B, marked node with 125I seed (n=32); and group C, marked node with clip and 125I seed (n=41). Dual tracers were used to search for SLNs after NAT. The main endpoint was the detection rate of marked nodes and false-negative rate (FNR). RESULTS: The detection rates using the TAD localization technique were 82.6% (28/34), 100% (32/32), and 100% (41/41) for groups A, B, and C, respectively (P>0.05). The FNR rates were 15.8%, 5.9%, and 5.6% among group A, B, and C, respectively (P>0.05). The FNR rates in cN1 patients were 5.1%, 2.7%, and 2.6%, among these three groups, respectively (P>0.05). The change in distance between 125I seeds and clips in axillary lymph nodes was <3 mm. The FNR rates of TAD guided by dye tracer, radiolabeled tracer, and dual tracers were 5.4%, 5.2%, and 3.4%, respectively (P>0.05). The negative predictive values were 93.0%, 93.0%, and 95.2%, respectively (P>0.05). CONCLUSION: Considering inexpensive and detect rate of 125I seeds, it is recommended that placement of 125I seeds to localize metastatic nodes in neoadjuvant setting. The TAD guided by dye tracer is also feasible for axillary de-escalation surgery after NAT in countries or regions without radiolabeled colloid.
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PURPOSE: To investigate the effects and mechanism of miRNA-153 on breast cancer cells in vitro and in vivo. MATERIAL AND METHODS: The cells and mice were divided into five groups: miRNA-153 mimic, miRNA-153 NC, miRNA-153 inhibitor, miRNA-153 inhibitor-NC, and blank control groups. The real-time PCR and western blot were used to detect the rictor expression regulated by miRNA-153. The western blot was used to explore the expression levels of p-Akt Ser473, p-SGK1 Ser422, and p-FOXO1 Thr24 regulated by miRNA-153. The H&E stain was used to detect the morphology and vitality of tumor cells. Flow cytometry analysis or TUNEL detection was used to evaluate the apoptosis of tumor cells. RESULTS: MiRNA-153 was significantly reduced in breast cancer cell lines. The real-time PCR and western blot assay suggested that the miRNA-153 downregulation of rictor expression, which was correlated with the antitumor effects both in vitro and in vivo. The western blot assay also showed that the expression levels of p-Akt Ser473, p-SGK1 Ser422, and p-FOXO1 Thr24 were largely reduced in miRNA-153 treated group, which indicated that miRNA-153 inhibited breast cancer growth by regulation of mTORC2 signaling pathway. The H&E stain demonstrated that the morphology and vitality of tumor cells in tumor tissues were influenced in miRNA-153 mimic treated group. The TUNEL detection also showed a great quantity of apoptotic cells in the miRNA-153 mimic group. CONCLUSIONS: All these results uncovering that the miRNA-153 inhibited breast cancer growth via regulation of mTORC2 signaling pathway, which provided breast cancer treatment a novel direction.
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MicroARNs , Neoplasias , Animales , Apoptosis/genética , Línea Celular Tumoral , Proliferación Celular/genética , Diana Mecanicista del Complejo 2 de la Rapamicina/genética , Diana Mecanicista del Complejo 2 de la Rapamicina/metabolismo , Ratones , MicroARNs/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de SeñalRESUMEN
Introduction: Breast cancer (BC) is associated with an inflammatory microenvironment. In BC, epidemiological evidence suggests that inflammation is associated with a poor prognosis. However, approaches to determine the extent of inflammation in the tumor microenvironment remain unclear. Material and methods: We downloaded the expression profiles and corresponding clinicopathological information of 1050 BC tissues and 59 cases of normal breast tissue from The Cancer Genome Atlas (TCGA) dataset. Similarly, data of 1050 BC tissues were downloaded from Gene Expression Omnibus (GEO) and 200 inflammation-related genes were downloaded from the MSigDB database. We developed an inflammatory risk model to reflect the immune microenvironment in BC. Results: Multivariate Cox analysis showed that the risk score was an independent predictor of overall survival (OS). Inflammatory signature was significantly associated with clinical and molecular features and could serve as an independent prognostic factor for BC patients. Furthermore, most immune cells were significantly less infiltrated in the high-risk group than in the low-risk group. There was a significant difference in survival time between the group with a high and low tumor mutational burden (TMB) score, and the survival time of the patients with a low TMB was significantly higher than that of the high-risk group. The risk scores were significantly lower in patients who responded to immunotherapy (complete response/partial response - CR/PR) than in patients who did not respond to immunotherapy (stable disease/progressive disease - SD/PD). Conclusions: We developed and validated an inflammatory risk model, which served as an independent prognostic indicator and reflected immune response intensity in the BC microenvironment.
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To explore the mechanism of TBX5 abnormal expression in simple congenital heart disease (CHD), 100 CHD venous blood, 50 CHD heart tissues, and 5 non-CHD heart tissues were involved in this study. The mutation and methylation in the 1 200 bp region upstream of TBX5 gene were detected by high-performance liquid chromatography (DHPLC) and methylation-sensitive restriction endonuclease (MS-RE), respectively. The binding site of NKX2-5 to Tbx5 predicted by P-MATCH software was validated by EMSA (Electrophoretic mobility shift assay). Tbx5 gene expression in mouse cardiac muscle cell H9C2(2-1) transfected with NKX2-5 expression vector was evaluated. No mutation was found in all patients. Both non-CHD and CHD heart tissues had the same methylation in the two CpG islands. Exogenous Nkx2-5 efficiently activated the transcription of the endogenous Tbx5 gene in H9C2 (2-1) cells. EMSA showed that the special binding band appeared when Nkx2-5 existed. These results indicates that the down expression of TBX5 might not be caused by mutation and methylation in the 1 200 bp region upstream of gene, and might be regulated by abnormal expression of NKX2-5 gene in heart muscle of CHD.
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Cardiopatías Congénitas/genética , Proteínas de Dominio T Box/fisiología , Animales , Islas de CpG/genética , Metilación de ADN/genética , Ensayo de Cambio de Movilidad Electroforética , Femenino , Humanos , Recién Nacido , Ratones , Embarazo , Regiones Promotoras Genéticas/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Proteínas de Dominio T Box/genéticaRESUMEN
BACKGROUND: Our previous research has suggested that genes around D12S1056 in 12q13 may confer susceptibility to ventricular septal defect (VSD) in humans. The present study was to define the chromosome region assignment by transmission disequilibrium test (TDT), and to identify the important candidate gene by family-based association study and haplotype analysis. METHODS: Surrounding D12S1056, ten microsatellite markers including D12S329, D12S305, D12S1662, D12S1056, D12S1293, D12S334, D12S102, D12S83, D12S1655 and D12S1691 were chosen, and TDT was performed in 62 nuclear family trios each consisting of an affected child and two healty parents. Subsequently, the GLI gene, a positional candidate gene that maps to the target region, was selected for further analysis. Three single nucleotide polymorphisms (SNPs), G11888C, G11388A, and G11625T, were selected for family-based association study and haplotype analysis. RESULTS: VSD was significantly associated with all selected markers except D12S1691 [72.2 centi morgen (cM)] and D12S1700 (75.76 cM). VSD was also significantly associated with G11888C (chi(2) = 5.918, P = 0.015), G11388A (chi(2) = 8.067, P = 0.005), and G11625T (chi(2) = 11.842, P = 0.001). Haplotype analysis showed a strong linkage disequilibrium between G11888C and G11388A (D' = 0.999), but in significant (chi(2) = 1.035, df = 2, P > 0.05). CONCLUSIONS: The susceptibility gene of VSD was mapped to 3.56 cM in 12q13 by TDT, and the GLI gene, an important candidate in the target region, was associated with VSD.
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Cromosomas Humanos Par 12 , Predisposición Genética a la Enfermedad , Defectos del Tabique Interventricular/genética , Factores de Transcripción/genética , Niño , Preescolar , Mapeo Cromosómico , Femenino , Haplotipos , Humanos , Desequilibrio de Ligamiento , Masculino , Repeticiones de Microsatélite , Proteína con Dedos de Zinc GLI1RESUMEN
To study the expression of TBX5, a key gene during heart formation, full-length TBX5 cDNA was cloned from Wistar rat embryonic heart (GenBank Accession No. AY859491). The cDNA and predicted amino acid sequences were different from those previously reported in GenBank. The expression profile of TBX5 in rat tissues was studied by RT-PCR and Northern blot. TBX5 was expressed in many rat tissues as a single transcript, and the highest level of expression was found in the heart. For subcellular localization, TBX5 was expressed as an EGFP fusion protein in rat hepatocarcinoma cells. Results showed that TBX5 was nuclear. In addition, TBX5-GST fusion protein was obtained by prokaryotic expression. These findings provide a good basis for further identification of TBX5-related transcription factors and protein-protein interaction studies among each other.
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ADN Complementario/genética , Regulación del Desarrollo de la Expresión Génica , Miocardio/metabolismo , Proteínas de Dominio T Box/genética , Animales , Secuencia de Bases , Núcleo Celular/metabolismo , Clonación Molecular , ADN Complementario/química , Exones , Vectores Genéticos , Glutatión Transferasa/genética , Glutatión Transferasa/metabolismo , Corazón/embriología , Intrones , Datos de Secuencia Molecular , Miocardio/citología , Ratas , Ratas Wistar , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Análisis de Secuencia de ADN , Proteínas de Dominio T Box/metabolismoRESUMEN
OBJECTIVE: In the candidate region 12q13 of simple congenital heart disease(CHD), four single nucleotide polymorphisms(SNPs) in HOXC4 gene were chosen in order to investigate the distribution of SNP and haplotypes in simple CHD patients and normal people. METHODS: The genotype of 4 SNPs in 108 simple CHD patients and 200 normal people were analyzed by restriction fragment length polymorphism(RFLP) and denaturing high-performance liquid chromatography(DHPLC). The statistical contingency table method was used to analyze SNP genotype frequency and gene frequency in patients and control group; then, the haplotypes were established and their frequencies in the two groups were assessed by PHASE software. RESULTS: C16476T polymorphism was not detected; A17860G located in 3' flanking sequence of HOXC5 gene displayed significant difference between the two groups. The G allele frequency in simple CHD patients was higher than that in healthy controls(P < 0.05); the distribution of frequencies of 4 haplotypes showed significant difference(P < 0.01). CONCLUSION: The A17860G located in 3'flanking sequence of HOXC5 gene is associated with simple CHD; the risk of CHD in the persons with G17860 is higher than that in those with A17860. the haplotype of 3 SNPs may be linked with the susceptible gene of simple CHD.
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Cromosomas Humanos Par 12/genética , Haplotipos/genética , Cardiopatías Congénitas/genética , Proteínas de Homeodominio/genética , Polimorfismo de Nucleótido Simple , Adolescente , Adulto , Alelos , Secuencia de Bases , Niño , Análisis Mutacional de ADN , Frecuencia de los Genes , Predisposición Genética a la Enfermedad/genética , Genotipo , Humanos , Persona de Mediana Edad , Datos de Secuencia Molecular , Adulto JovenRESUMEN
AIM: To investigate the expression of c-myc target from laryngeal cancer cells (MTLC) gene in gastric carcinoma (GC) tissues and the effect of MTLC over-expression on gastric carcinoma cell line BGC823. METHODS: RT-PCR was performed to determine the expression of MTLC mRNA in GC and matched control tissues. BGC823 cells were transfected with an expression vector pcDNA3.1-MTLC by liposome and screened by G418. Growth of cells expressing MTLC was observed daily by manual counting. Apoptotic cells were determined by TdT-mediated dUTP nick-end labeling (TUNEL) assay. RESULTS: The expression of MTLC mRNAs was down-regulated in 9(60%) of 15 cases of GC tissues. The growth rates of the BGC823 cells expressing MTLC were indistinguishable from that of control cells. A marked acceleration of apoptosis was observed in MTLC-expressing cells. CONCLUSION: MTLC was down-regulated in the majority of GC tissues and could promote apoptosis of GC cell lines, which suggests that MTLC may play an important role in the carcinogenesis of gastric carcinoma.
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Regulación Neoplásica de la Expresión Génica , Proteínas Nucleares/genética , Neoplasias Gástricas/fisiopatología , Apoptosis/fisiología , División Celular/fisiología , Línea Celular Tumoral , Regulación hacia Abajo/fisiología , Humanos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Neoplasias Gástricas/patología , TransfecciónRESUMEN
OBJECTIVE: To analyze the genetic polymorphism of 6 short tandem repeat (STR) loci on chromosome 7p14-15 and 8 STR loci on chromosome 12q13 in Chinese north Hans. METHODS: Fluorescence-labeling polymerase chain reaction and capillary electrophoresis were used to analyze the genetic polymorphism of 100 randomly selected individuals from Chinese north Han nationality at 6 STR loci (D7S1808, D7S2250, D7S2251, D7S683, D7S656 and D7S528) on chromosome 7p14-15 and 8 STR loci(D12S1056, D12S1293, D12S83, D12S1655, D12S1662, D12S334, D12S137 and D12S102) on chromosome 12q13. RESULTS: In the Chinese north Han population, 7 alleles and 24 genotypes, 8 alleles and 27 genotypes, 7 alleles and 22 genotypes, 4 alleles and 10 genotypes, 6 alleles and 17 genotypes, 5 alleles and 13 genotypes were observed at D7S1808, D7S2250, D7S2251, D7S683, D7S656 and D7S528. The heterozygosities at the above 6 STR loci were 86%, 88%, 83%, 79%, 85% and 80%, respectively. Five alleles and 15 genotypes, 5 alleles and 15 genotypes, 8 alleles and 29 genotypes, 6 alleles and 17 genotypes, 6 alleles and 17 genotypes, 6 alleles and 19 genotypes, 5 alleles and 13 genotypes, 7 alleles and 24 genotypes were observed at D12S1056, D12S1293, D12S83, D12S1655, D12S1662, D12S334, D12S137 and D12S102. The heterozygosities at the above 8 STR loci were 86%, 84%, 87%, 82%, 84%, 85%, 81% and 89%, respectively. CONCLUSION: The distributions of allele frequencies of 6 STR loci on chromosome 7p14-15 and of 8 STR loci on chromosome 12q13 were consistent with the Hardy-Weinberg equilibrium. The highly genetic polymorphism was observed in Chinese north Han population.
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Cromosomas Humanos Par 12/genética , Cromosomas Humanos Par 7/genética , Repeticiones de Microsatélite/genética , Polimorfismo Genético/genética , Pueblo Asiatico/genética , China , Humanos , Reacción en Cadena de la PolimerasaRESUMEN
The mechanism of telomerase activation is still not clear till now. In order to understand the expressional mode of telomerase and the mechanism of telomerase activation in laryngeal carcinogenesis, we cloned a fragment of hTERT cDNA and prepared a monoclonal antibody against hTERT. We performed immuno-histochemical staining in laryngeal cancer tissues using this antibody. We found that the frequencies of hTERT positive cells were positively correlated with undifferentiation of cancer tissues and that the expression of hTERT was positively correlated with levels of c-Myc, which suggested that c-Myc might play an important role in activation of telomerase. These results revealed that overexpression of c-Myc upregulated telomerase, which in turn resulted in immortalization of laryngeal squamous cells, and this mechanism existed not only in the initiation of laryngeal carcinogenesis but also in the whole process of cancer development.
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Anticuerpos Monoclonales/inmunología , ADN Complementario/genética , Neoplasias Laríngeas/patología , Telomerasa/genética , Clonación Molecular , Proteínas de Unión al ADN , Vectores Genéticos/genética , Glutatión Transferasa/genética , Glutatión Transferasa/metabolismo , Humanos , Inmunohistoquímica , Neoplasias Laríngeas/genética , Neoplasias Laríngeas/inmunología , Proteínas Proto-Oncogénicas c-myc/inmunología , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Telomerasa/inmunología , Telomerasa/metabolismoRESUMEN
OBJECTIVE: To identify and characterize laryngeal cancer related novel genes located on chromosome 6q25. METHODS: Electric hybridization was performed in human genome database using EST (expression sequence tag) as probe. Novel genes were deduced by software from positive DNA clones and their cDNAs were amplified by RT-PCR using primers designed according to the sequence of the putative genes. RESULTS: A novel gene was cloned successfully. The full length of this gene was about 21 kb. It contained two exons and produced a 1006 bp transcript coding a protein with 235 amino acid residues. It's 5'flanking sequence contained two binding sites of oncoprotein c-Myc, thus it was named MTLC (c-Myc target from laryngeal cancer cells). Homologous assay showed that MTLC exhibited little overall homology to known human proteins but it exhibited good overall homology to mouse MT-MC1 protein with an identity of 78%. The primary structure of MTLC protein contained a nuclear location signal motif, but it did not have other conserved domains. The results of subcellular location experiment showed that MTLC expressed in nuclei of human hepatocellular carcinoma cell line Bel7402 cells, while a wide distribution of MTLC in various tissues was demonstrated by Northern blotting. CONCLUSION: MTLC may play an important role as a target gene of c-Myc and as a transcription factor in keeping the normal physiological process of cells.
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Cromosomas Humanos Par 6/genética , Neoplasias Laríngeas/genética , Proteínas Nucleares/genética , Secuencia de Aminoácidos , Secuencia de Bases , Northern Blotting , Clonación Molecular , ADN Complementario/química , ADN Complementario/genética , Expresión Génica , Proteínas Fluorescentes Verdes , Humanos , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Microscopía Fluorescente , Datos de Secuencia Molecular , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Alineación de Secuencia , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Células Tumorales CultivadasRESUMEN
This work is to investigate the mutation and expression of TBX5 gene in human simple congenital heart disease. The mutations of eight exons of TBX5 gene in 61 CHD family members (a total of 216 individuals including 65 patients and 151 normal relatives) were examined by PCR-DGGE. Using beta-actin as internal control, the differential expression between 34 myocardium samples from simple congenital heart disease patients and three normal controls was conducted by RT-PCR. There is no mutation detected in all samples; The mRNA expression levels of TBX5 gene show descent tendency in samples of simple congenital heart disease compared with normal controls. The mutations in coding region of TBX5 gene do not cause human simple congenital heart disease, but the abnormality in transcription level of TBX5 gene maybe a kind of mechanism causing human simple congenital heart disease.