Objective:To investigate the dynamic changes of cellular miRNA expression profiles in EBV latently infected Raji cells upon reactivation with Phorbol ester (TPA).Methods:Total RNA was extracted using TRIzol reagent from Raji cells treated with TPA at different time points (0 h, 24 h, 48 h). Small RNA libraries were constructed and sequenced on an Illumina SE50 platform. Differentially expressed miRNAs were identified, and their target genes were predicted and functionally annotated. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were carried out through online tools. Additionally, miRanda and RNAhybrid software were used to predict cellular miRNAs targeting the EBV genome. Real-time RT-qPCR was employed to validate the expression levels of differentially expressed novel miRNAs.Results:High-throughput sequencing identified 1 301 celluar miRNAs, comprising 1 189 known and 112 novel miRNAs. A total of 264 known differentially expressed cellular miRNAs and 13 novel miRNAs were identified through high-throughput miRNA sequencing. Secondary structure prediction revealed that the novel miRNAs exhibited typical pre-miRNA hairpin structures. Stem-loop quantitative real-time PCR (RT-qPCR) validation of Novel_miR_183 and Novel_miR_242 did not exhibit a statistically significant difference ( F=1.407, P=0.370 7 for Novel_miR_183; F=1.277, P=0.397 0 for Novel_miR_242) between the TPA-stimulated and untreated groups. Target gene prediction analysis revealed that the differentially expressed cellular miRNAs were involved in various important biological processes and signaling pathways. Furthermore, 1 189 known cellular miRNAs and 108 novel miRNAs were predicted to target the EBV genome. Conclusions:Treatment of Raji cells with TPA stimulation successfully reactivated Raji cells and significantly altered their miRNA expression patterns. Differentially expressed miRNAs were identified, suggesting that these miRNAs probably play crucial roles in regulating EBV infection and replication by directly targeting the EBV genome.

Objective:To establish a protocol for virus isolation using the mixed method, and evaluate the efficacy of the suspended method and the mixed method in isolating herpes simplex virus (HSV).Methods:Simulated HSV-infected clinical samples were prepared using HSV-1 F strain and CDC-P1 strain. Both the suspended method and the mixed method were used to isolate HSV-1 from these samples. The virus isolation efficiency of the mixed method under various conditions was assessed. These conditions included different multiplicity of infection (MOI), cell seeding densities, and virus adsorption times.The 50% tissue culture infective dose (TCID 50) assay was used for the assessment. The positive rates of virus detection under low viral load conditions were compared between the two methods. Results:Under the conditions of a MOI of 0.005, a virus adsorption time of 15 min, and a cell seeding density of 1×10 6 cell/ml, the mixed method achieved effective isolation of HSV-1. When the virus titer of the sample was 100 TCID 50/ml, the positivity rate of the mixed method reached 100.0%, while the positivity rates of the suspended method were 50.7% (38/75) and 52.0% (39/75) after cultured for 72 h and 96 h, respectively. When the virus titer of the sample was 10 TCID 50/ml, the positivity rate of the mixed method was 100.0%, while the positivity rate of the suspension method was 0. Conclusions:The mixed method exhibits significantly higher efficiency in HSV isolation compared with the suspended method. Under the conditions of high viral load, both the suspended method and the mixed method can be effective in isolating HSV-1. For clinical samples with low viral loads, the mixed method has greater applicability.

Objective:To explore the safety and efficacy of computed tomography (CT) guided percutaneous interstitial brachytherapy in the treatment of recurrent cervical cancer with isolated lesions in the radiated field.Methods:A retrospective analysis was conducted on the clinical data of 30 patients with recurrent cervical cancer with isolated lesions in the radiated field who underwent CT guided percutaneous interstitial implantation for close range radiation treatment at Zhengzhou University Affiliated Cancer Hospital from March 2023 to August 2024. Under local anesthesia, a needle was implanted into the recurrent tumor in the pelvic or abdominal wall of the patients percutaneously guided by CT. The target area was delineated to ensure full dose coverage. The prescribed dose for high-risk clinical target areas was 600 cGy/time, once a week, followed by close range radiotherapy. The number of implanted needles were recorded, and the target area, radiation dose, and other parameters were evaluated through dose volume parameter maps. The degree of lesion shrinkage and the occurrence of complications during and after treatment were observed.Results:30 patients underwent a total of 72 rounds of brachytherapy with implantation, with a technical success rate of 100% (72/72). 20 cases received 2 treatments, 8 cases received 3 treatments, and 2 cases received 4 treatments; 4 cases used 1needle, 20 cases used 2 needles, 4 cases used 3 needles, and 2 cases used 4 needles. The high-risk clinical target dose D 90 was (718.17±222.61) cGy. The average dose D 2cc of 2 cm 3 surrounding the bladder, rectum, sigmoid colon, and small intestine was (168.29±53.80) cGy, (178.87±105.38) cGy, (136.05±78.06) cGy, and (288.91±117.49) cGy, respectively. The median follow-up time was 11 months. Among the 30 patients, there were 12 cases of complete remission,14 cases of partial remission, 3 cases of stable disease, and 1 case of disease progression, with an objective remission rate of 86.7%. None of the patients experienced significant bleeding or pain during treatment. After treatment, 3 patients with recurrent lymph nodes near the rectum developed grade 1 radiation proctitis, which was remitted after treatment. No significant complications were observed in the remaining patients. Conclusion:CT guided percutaneous brachytherapy is safe and feasible for the recurrence of single lesions in the radiated field of cervical cancer.

Objective:To investigate the prognostic impact of perineural invasion in patients with stageⅢ colon cancer and to clarify its guidance value for the duration of postoperative adjuvant chemotherapy.Methods:This study employed a retrospective cohort study method. It analyzed 426 patients with stageⅢ colon cancer who underwent radical surgery at Sun Yat-sen University Cancer Center and Longyan First Affiliated Hospital of Fujian Medical University, between April 2008 and June 2020. Inclusion criteria: patients received at least 3 months of adjuvant CapeOX therapy post-surgery, had complete pathological data, and were followed up for at least 12 months after the last chemotherapy. Among these patients, 231 were male, the median age was 59 (50~67) years, and 263 tumors were located in the right-sided colon. Postoperative pathology indicated that 107 cases (25.12%) had neural invasion, and 131 patients (30.75%) had vascular tumor thrombus. All patients received at least 4 cycles of postoperative CapeOX adjuvant chemotherapy, with 193 patients receiving 8 cycles and 233 patients receiving 4 to 7 cycles of adjuvant chemotherapy. The study analyzed the impact of neural invasion status and the duration of adjuvant chemotherapy on disease-free survival (DFS). Furthermore, within subgroups stratified by different risk levels (referencing the criteria proposed by the IDEA study: high risk: T4, N2 or T4N2; low risk: T3N1) and different neural invasion statuses, the impact of the duration of adjuvant chemotherapy on prognosis was analyzed.Results:The median follow-up time for the entire cohort was 94.00 months (55.27-128.80 months). Multivariate Cox analysis indicated that pathological T stage T4 (HR = 2.457, 95%CI: 1.499-4.029, P<0.001) and postoperative pathological confirmation of perineural invasion (HR = 2.465, 95% CI: 1.519-4.000, P<0.001) were independent adverse prognostic factors for 5-year DFS. In the perineural invasion-positive group, the 5-year DFS for patients who received 8 cycles of postoperative adjuvant CapeOX chemotherapy was 86.90%, compared to 58.22% for those who received 4-7 cycles, with statistically significant differences (both P<0.05). In the perineural invasion-negative group, the 5-year DFS for patients who received 8 cycles was 88.66%, compared to 90.99% for those who received 4-7 cycles, with no statistically significant differences ( P=0.929). Among IDEA high-risk patients with perineural invasion, the 5-year DFS was 91.81% for those who received 8 cycles versus 50.66% for those who received 4-7 cycles, showing a statistically significant difference ( P=0.003). In IDEA high-risk patients without perineural invasion, the 5-year DFS for those who received 8 cycles was 82.28% compared to 87.32% for those who received 4-7 cycles, with no statistically significant difference ( P=0.806). In the IDEA low-risk patients, no differences were observed in the 5-year DFS between patients receiving 8 cycles and those receiving 4-7 cycles of adjuvant CapeOX chemotherapy in both perineural invasion-positive and negative subgroups (both P>0.05). Conclusion:Perineural invasion serves as a significant prognostic factor for 5-year DFS in stage Ⅲ colon cancer patients who have undergone radical surgery and postoperative adjuvant chemotherapy. It can also be considered an important reference factor in deciding the duration of postoperative adjuvant chemotherapy.

With the continuous development of imaging techniques such as magnetic resonance imaging,the detection rate of brain metastases is increasing.Although the incidence rate of cystic brain metastasis is far lower than that of solid brain metastasis,patients with cystic brain metastasis are in urgent condition and have obvious space occupying effect,which is an urgent clinical problem.Previous literature has reported that cystic brain metastasis is more common in breast cancer and lung adenocarcinoma,especially in lung cancer patients with positive driver gene.This article reports a case of small cell lung cancer with cystic brain metastasis,which started with neurological symptoms,and was clinically cured under a multidisciplinary diagnosis and treatment model.Through dynamic imaging evaluation and molecular residual lesion detection,the patient can avoid overtreatment and achieved a relatively higher quality of life on the basis of prolonging survival.

Objective To investigate the correlation between G protein-coupled receptor family C group 6 member A(GPRC6A)gene polymorphisms and their expression and pulmonary infections in elderly patients with chronic heart failure(CHF).Methods 138 elderly CHF patients admitted to the Xianyang First People's Hospital from January 2021 to January 2024 were selected as the research subjects,and were divided into an infected group(n=42)and an uninfected group(n=96)based on their lung infection status.Polymerase chain reaction(PCR)was used to detect polymorphisms at the rs6901250 and rs1606365 loci of the GPRC6A gene.The allele and genotype frequency distributions of the infected and uninfected groups were compared.Logistic regression modeling was used to analyze the s6901250 and rs1606365 loci under three genetic models(co-dominant,dominant and reces-sive)and lung infections in elderly patients with CHF.Real-time fluorescence quantitative PCR(RT-qPCR)method was used to detect the expression level of GPRC6A gene.The predictive value of the mRNA expression level of the GPRC6A gene for the development of pulmonary infections in elderly patients with CHF was analyzed by applying the receiver operator characteristic(ROC)curve.Results The distribution of genotypes at loci rs6901250 and rs1606365 of the GPRC6A gene in both the infected and uninfected groups of the lungs of elderly CHF patients conformed to the Hardy-Weinberg equilibrium law(χ2=0.199～0.376,all P>0.05),which was representative of the population.Compared with the uninfected group,the frequency of allele A at locus rs6901250(57.14%vs 41.67%)was significantly higher in the infected group,Allele G(54.76%vs.37.50%)and genotype GG(14.06%vs 29.99%)frequencies were significantly higher at locus rs1606365,and the differences were statistically significant(χ2=5.628,7.114,6.849,all P<0.05).At locus rs6901250,in the co-dominant model(GG vs AA)and the dominant model(GA+AA vs GG),the elderly CHF patients with AA genotype the risk of lung infection was higher than that of GG genotype(OR=1.753,1.546,all P<0.05);.rs1606365 locus showed that the risk of lung infection was higher than that of CC genotype in el-derly CHF patients with GG genotype under all three genetic models of co-dominant model(CC vs GG),dominant model(CG+GG vs CC)and recessive model(CG+CC vs GG)(OR=1.833,1.741,0.695,all P<0.05).The mRNA expression level of GPR-C6A gene in the lung-infected group of elderly CHF patients(1.43±0.35)was significantly higher than that in the uninfected group(1.02±0.21),and the difference was statistically significant(t=8.515,P<0.001).The results of the ROC curve analysis showed that the GPRC6A gene expression level predicted lung infection in elderly CHF patients with an AUC value of 0.895,a cut-offvalue of 1.37,and sensitivity and specificity of 85.7%and 66.7%,respectively.Conclusion The AA genotype at the rs6901250 locus and the GG genotype at the rs1606365 locus of the GPRC6A gene increased the risk of developing lung infec-tions in elderly patients with CHF.MRNA expression levels of the GPRC6A gene were elevated in the infected group,and its ex-pression level could be used as a predictive indicator for the development of lung infections in elderly patients with CHF.

Objective To investigate the correlation between G protein-coupled receptor family C group 6 member A(GPRC6A)gene polymorphisms and their expression and pulmonary infections in elderly patients with chronic heart failure(CHF).Methods 138 elderly CHF patients admitted to the Xianyang First People's Hospital from January 2021 to January 2024 were selected as the research subjects,and were divided into an infected group(n=42)and an uninfected group(n=96)based on their lung infection status.Polymerase chain reaction(PCR)was used to detect polymorphisms at the rs6901250 and rs1606365 loci of the GPRC6A gene.The allele and genotype frequency distributions of the infected and uninfected groups were compared.Logistic regression modeling was used to analyze the s6901250 and rs1606365 loci under three genetic models(co-dominant,dominant and reces-sive)and lung infections in elderly patients with CHF.Real-time fluorescence quantitative PCR(RT-qPCR)method was used to detect the expression level of GPRC6A gene.The predictive value of the mRNA expression level of the GPRC6A gene for the development of pulmonary infections in elderly patients with CHF was analyzed by applying the receiver operator characteristic(ROC)curve.Results The distribution of genotypes at loci rs6901250 and rs1606365 of the GPRC6A gene in both the infected and uninfected groups of the lungs of elderly CHF patients conformed to the Hardy-Weinberg equilibrium law(χ2=0.199～0.376,all P>0.05),which was representative of the population.Compared with the uninfected group,the frequency of allele A at locus rs6901250(57.14%vs 41.67%)was significantly higher in the infected group,Allele G(54.76%vs.37.50%)and genotype GG(14.06%vs 29.99%)frequencies were significantly higher at locus rs1606365,and the differences were statistically significant(χ2=5.628,7.114,6.849,all P<0.05).At locus rs6901250,in the co-dominant model(GG vs AA)and the dominant model(GA+AA vs GG),the elderly CHF patients with AA genotype the risk of lung infection was higher than that of GG genotype(OR=1.753,1.546,all P<0.05);.rs1606365 locus showed that the risk of lung infection was higher than that of CC genotype in el-derly CHF patients with GG genotype under all three genetic models of co-dominant model(CC vs GG),dominant model(CG+GG vs CC)and recessive model(CG+CC vs GG)(OR=1.833,1.741,0.695,all P<0.05).The mRNA expression level of GPR-C6A gene in the lung-infected group of elderly CHF patients(1.43±0.35)was significantly higher than that in the uninfected group(1.02±0.21),and the difference was statistically significant(t=8.515,P<0.001).The results of the ROC curve analysis showed that the GPRC6A gene expression level predicted lung infection in elderly CHF patients with an AUC value of 0.895,a cut-offvalue of 1.37,and sensitivity and specificity of 85.7%and 66.7%,respectively.Conclusion The AA genotype at the rs6901250 locus and the GG genotype at the rs1606365 locus of the GPRC6A gene increased the risk of developing lung infec-tions in elderly patients with CHF.MRNA expression levels of the GPRC6A gene were elevated in the infected group,and its ex-pression level could be used as a predictive indicator for the development of lung infections in elderly patients with CHF.

Objective:To investigate the prognostic impact of perineural invasion in patients with stageⅢ colon cancer and to clarify its guidance value for the duration of postoperative adjuvant chemotherapy.Methods:This study employed a retrospective cohort study method. It analyzed 426 patients with stageⅢ colon cancer who underwent radical surgery at Sun Yat-sen University Cancer Center and Longyan First Affiliated Hospital of Fujian Medical University, between April 2008 and June 2020. Inclusion criteria: patients received at least 3 months of adjuvant CapeOX therapy post-surgery, had complete pathological data, and were followed up for at least 12 months after the last chemotherapy. Among these patients, 231 were male, the median age was 59 (50~67) years, and 263 tumors were located in the right-sided colon. Postoperative pathology indicated that 107 cases (25.12%) had neural invasion, and 131 patients (30.75%) had vascular tumor thrombus. All patients received at least 4 cycles of postoperative CapeOX adjuvant chemotherapy, with 193 patients receiving 8 cycles and 233 patients receiving 4 to 7 cycles of adjuvant chemotherapy. The study analyzed the impact of neural invasion status and the duration of adjuvant chemotherapy on disease-free survival (DFS). Furthermore, within subgroups stratified by different risk levels (referencing the criteria proposed by the IDEA study: high risk: T4, N2 or T4N2; low risk: T3N1) and different neural invasion statuses, the impact of the duration of adjuvant chemotherapy on prognosis was analyzed.Results:The median follow-up time for the entire cohort was 94.00 months (55.27-128.80 months). Multivariate Cox analysis indicated that pathological T stage T4 (HR = 2.457, 95%CI: 1.499-4.029, P<0.001) and postoperative pathological confirmation of perineural invasion (HR = 2.465, 95% CI: 1.519-4.000, P<0.001) were independent adverse prognostic factors for 5-year DFS. In the perineural invasion-positive group, the 5-year DFS for patients who received 8 cycles of postoperative adjuvant CapeOX chemotherapy was 86.90%, compared to 58.22% for those who received 4-7 cycles, with statistically significant differences (both P<0.05). In the perineural invasion-negative group, the 5-year DFS for patients who received 8 cycles was 88.66%, compared to 90.99% for those who received 4-7 cycles, with no statistically significant differences ( P=0.929). Among IDEA high-risk patients with perineural invasion, the 5-year DFS was 91.81% for those who received 8 cycles versus 50.66% for those who received 4-7 cycles, showing a statistically significant difference ( P=0.003). In IDEA high-risk patients without perineural invasion, the 5-year DFS for those who received 8 cycles was 82.28% compared to 87.32% for those who received 4-7 cycles, with no statistically significant difference ( P=0.806). In the IDEA low-risk patients, no differences were observed in the 5-year DFS between patients receiving 8 cycles and those receiving 4-7 cycles of adjuvant CapeOX chemotherapy in both perineural invasion-positive and negative subgroups (both P>0.05). Conclusion:Perineural invasion serves as a significant prognostic factor for 5-year DFS in stage Ⅲ colon cancer patients who have undergone radical surgery and postoperative adjuvant chemotherapy. It can also be considered an important reference factor in deciding the duration of postoperative adjuvant chemotherapy.

With the continuous development of imaging techniques such as magnetic resonance imaging,the detection rate of brain metastases is increasing.Although the incidence rate of cystic brain metastasis is far lower than that of solid brain metastasis,patients with cystic brain metastasis are in urgent condition and have obvious space occupying effect,which is an urgent clinical problem.Previous literature has reported that cystic brain metastasis is more common in breast cancer and lung adenocarcinoma,especially in lung cancer patients with positive driver gene.This article reports a case of small cell lung cancer with cystic brain metastasis,which started with neurological symptoms,and was clinically cured under a multidisciplinary diagnosis and treatment model.Through dynamic imaging evaluation and molecular residual lesion detection,the patient can avoid overtreatment and achieved a relatively higher quality of life on the basis of prolonging survival.

Objective:To establish a protocol for virus isolation using the mixed method, and evaluate the efficacy of the suspended method and the mixed method in isolating herpes simplex virus (HSV).Methods:Simulated HSV-infected clinical samples were prepared using HSV-1 F strain and CDC-P1 strain. Both the suspended method and the mixed method were used to isolate HSV-1 from these samples. The virus isolation efficiency of the mixed method under various conditions was assessed. These conditions included different multiplicity of infection (MOI), cell seeding densities, and virus adsorption times.The 50% tissue culture infective dose (TCID 50) assay was used for the assessment. The positive rates of virus detection under low viral load conditions were compared between the two methods. Results:Under the conditions of a MOI of 0.005, a virus adsorption time of 15 min, and a cell seeding density of 1×10 6 cell/ml, the mixed method achieved effective isolation of HSV-1. When the virus titer of the sample was 100 TCID 50/ml, the positivity rate of the mixed method reached 100.0%, while the positivity rates of the suspended method were 50.7% (38/75) and 52.0% (39/75) after cultured for 72 h and 96 h, respectively. When the virus titer of the sample was 10 TCID 50/ml, the positivity rate of the mixed method was 100.0%, while the positivity rate of the suspension method was 0. Conclusions:The mixed method exhibits significantly higher efficiency in HSV isolation compared with the suspended method. Under the conditions of high viral load, both the suspended method and the mixed method can be effective in isolating HSV-1. For clinical samples with low viral loads, the mixed method has greater applicability.