Feasibility analysis of rapid gene detection using intraoperative frozen tissues: comparison of intraoperative frozen tissues with paraffin-embedded tissues in epidermal growth factor receptor gene mutation detection of lung adenocarcinoma.

BACKGROUND: Epidermal growth factor receptor (EGFR) is the driver gene with the highest frequency of mutations in lung adenocarcinoma and can guide the development of targeted therapies. The detection of routine gene mutations must be performed after the preparation of paraffin samples in a standard polymerase chain reaction (PCR) laboratory, which is time-consuming. The Idylla™ EGFR fully automatic PCR system for rapid detection requires no special detection environment and completes the process in only 2.5 h. It has been applied to tissues embedded in paraffin. METHODS: The Idylla™ EGFR automated PCR system was used to detect EGFR gene mutations in intraoperative frozen fresh tissues and paraffin-embedded tissues from 47 enrolled patients with lung adenocarcinoma. The gold standard amplification refractory mutation system (ARMS) method for gene mutation detection was used for verification, and the concordance between the three detection results was compared, to investigate the feasibility of detecting rapid gene mutations in intraoperative frozen samples. RESULTS: The EGFR mutation rate in 47 fresh samples of lung adenocarcinoma was 61.7% (29/47), which is consistent with the mutation level of lung adenocarcinoma in the Asian population (38.8-64.0%). The concordance rate between the Idylla™ frozen tissues and paraffin-embedded tissues was 91.4% (43/47) when compared to the ARMS method, while the coincidence rate between the two methods was 93.6% (44/47). The three methods had a total consistency rate of 89.4% (42/47). CONCLUSIONS: The Idylla™ EGFR fully automatic PCR system directly detects EGFR mutations in fresh tissues. The operation is simple, the detection time is short, and the accuracy is high. The detection time is reduced to 1/4-1/3 of the original time while meeting clinical standards for detecting the gene status of patients, thus saving crucial time for individualized and accurate treatment of patients. The method has promising clinical application prospects.

A comparison of capillary electrophoresis and next-generation sequencing in the detection of immunoglobulin heavy chain H and light chain κ gene rearrangements in the diagnosis of classic hodgkin's lymphoma.

This study aimed to compare the application value of capillary electrophoresis and next-generation sequencing for immunoglobulin (IG) gene rearrangement in the diagnosis of classic Hodgkin's lymphoma. Twenty paraffin-embedded specimens from patients with classic Hodgkin's lymphoma were screened. For gene rearrangement detection, the ABI 3500 Genetic Analyzer and ABI Ion GeneStudio S5 Plus sequencing system were used, respectively, and the results were compared. Five cases with monoclonal rearrangements (25%, 5/20) were detected by Capillary Electrophoresis, and positivity for the FR1, FR2, FR3, and IGк loci was 5%, 10%, 10%, and 15%, respectively; 12 cases with monoclonal rearrangements (60%, 12/20) were detected by Next-generation Sequencing where the positivity of the above corresponding loci were 35%, 45%, 50%, and 30%, respectively. Among the 20 samples, 6 IGк clonal rearrangements were detected, and the usage frequency (66.7%) of IGкJ4 was the highest in the IGкJ subgroup. The usage frequency of IGкV1 and IGкV3 in the GкV sub-group was 33.3% and 33.3%, respectively. Twelve immunoglobulin heavy chain (IGH) clonal rearrangements were detected among the 20 samples, and the order of usage frequency in the IGH joining region J (IGHJ) subgroup was IGHJ4 > IGHJ5 > IGHJ6 > IGHJ3. The gene with the highest usage frequency in the IGH variable (IGHV) subgroup was IGHV3 (50%) and the percentage of IGHV mutations ranged from 0% ± 11.45% with an average frequency of 3.34%. Compared with Capillary Electrophoresis, Next-generation Sequencing showed a higher positivity in the detection of gene clonal rearrangements, was more accurate in the interpretation of results.

A retrospective analysis of the clinicopathological characteristics of large cell carcinoma of the lung.

The aim of the present study was to analyze and summarize the clinicopathological characteristics of large-cell lung carcinoma (LCLC) of the lung, in order to improve the definite diagnosis rate of LCLC. Clinicopathological data of 174 patients with LCLC, confirmed pathologically, were retrospectively reviewed. The 174 cases of LCLC accounted for 5.7% of the total lung cancer cases during the corresponding time period at the Affiliated Cancer Hospital of Tianjin Medical University (Tianjin, China), among which there were 131 males and 43 females with an average age of 61.4 years. The postoperative pathological diagnosis of the 174 cases showed 80 cases of classic LCLC, 64 cases of large cell neuroendocrine carcinoma (LCNEC), six cases of combined LCNEC, 19 cases of basaloid carcinoma, three cases of clear cell carcinoma and two cases of lymphoepithelioma-like carcinoma. Of the total 174 LCLC cases, 96 patients exhibited lymph node metastasis. LCLC is a highly aggressive malignancy with a high tendency of invasion and metastasis, although the incidence rate is low. A definite diagnosis of LCLC primarily relies on the pathological diagnosis. Each subtype of LCLC has its own pathomorphological and immunohistochemical characteristics.

Dopaminergic neuron-like cells derived from bone marrow mesenchymal stem cells by Lmx1α and neurturin overexpression for autologous cytotherapy in hemiparkinsonian rhesus monkeys.

Bone marrow-derived mesenchymal stem cells hold great potential for cytotherapeutics of neurodegenerative disorders, including Parkinson's disease. The neurotrophic factor neurturin can rescue dopaminergic neurons damaged during the disease process. Lmx1α can promote mesencephalic dopaminergic differentiation during embryogenesis. In this study, we tested a cytotherapeutic strategy combining NTN/Lmx1α gene therapy and cell transplantation to ameliorate disease progression in hemiparkinsonian rhesus. Rhesus BMSCs were prepared for autologous grafting by transfection with recombinant adenoviral vectors expressing secreted NTN and Lmx1α,and cultured in the presence of induce factors, particularly the Lmx1α regulatory factor sonic hedgehog, to guide dopaminergic differentiation. These induced rh-BMSCs exhibited gene/protein expression phenotypes resembling nigral dopaminergic neurons. They survived and retained dopaminergic function following stereotaxic injection into the MPTP-lesioned right-side substantia nigra as indicated by SPECT measurement of DAT activity. Injected cells preserved and supplemented the remaining endogenous population of dopamine neurons (TH-positive cell ipsilateral/contralateral ratio was 56.81% ± 7.28% vs. 3.86%±1.22% in vehicle-injected controls; p<0.05). Cell injection also partially restored motor function and reduce apomorphine-evoked rotation (p<0.05). Moreover, function recovery occurred earlier than in previous studies on injected BMSCs. Our findings demonstrate a promising strategy for restoration of PD-associated motor dysfunction by transplantation of autologous BMSCs overexpressing NTN/Lmx1α.