Heterozygous/dispermic complete mole confers a significantly higher risk for post-molar gestational trophoblastic disease.

Hydatidiform moles are classified at the genetic level as androgenetic complete mole and diandric-monogynic partial mole. Conflicting data exist whether heterozygous complete moles are more aggressive clinically than homozygous complete moles. We investigated clinical outcome in a large cohort of hydatidiform moles in Chinese patients with an emphasis on genotypical correlation with post-molar gestational trophoblastic disease. Consecutive products of conceptions undergoing DNA genotyping and p57 immunohistochemistry to rule out molar gestations were included from a 5-year period at Beijing Obstetrics and Gynecology Hospital. Patient demographics and clinical follow-up information were obtained. Post-molar gestational trophoblastic disease or gestational trophoblastic neoplasia was determined by the 2002 WHO/FIGO criteria. A total of 1245 products of conceptions were classified based on genotyping results into 219 complete moles, 250 partial moles, and 776 non-molar gestations. Among 219 complete moles, 186 were homozygous/monospermic and 33 were heterozygous/dispermic. Among 250 partial moles, 246 were triploid dispermic, 2 were triploid monospermic, and 2 were tetraploid heterozygous partial moles. Among 776 non-molar gestations, 644 were diploid without chromosomal aneuploidies detectable by STR genotyping and 132 had various genetic abnormalities including 122 cases of various trisomies, 2 triploid digynic-monoandric non-molar gestations, 7 cases of possible chromosomal monosomy or uniparental disomy. Successful follow-up was achieved in 165 complete moles: post-molar gestational trophoblastic disease developed in 11.6% (16/138 cases) of homozygous complete moles and 37.0% (10/27 cases) of heterozygous complete moles. The difference between the two groups was highly significant (p = 0.0009, chi-square). None of the 218 partial moles and 367 non-molar gestations developed post-molar gestational trophoblastic disease. In conclusion, heterozygous/dispermic complete moles are clinically more aggressive with a significantly higher risk for development of post-molar gestational trophoblastic disease compared with homozygous/monospermic complete moles. Therefore, precise genotyping classification of complete moles is important for clinical prognosis and patient management.

Measuring entanglement entropy and its topological signature for phononic systems.

Entanglement entropy is a fundamental concept with rising importance in various fields ranging from quantum information science, black holes to materials science. In complex materials and systems, entanglement entropy provides insight into the collective degrees of freedom that underlie the systems' complex behaviours. As well-known predictions, the entanglement entropy exhibits area laws for systems with gapped excitations, whereas it follows the Gioev-Klich-Widom scaling law in gapless fermion systems. However, many of these fundamental predictions have not yet been confirmed in experiments due to the difficulties in measuring entanglement entropy in physical systems. Here, we report the experimental verification of the above predictions by probing the nonlocal correlations in phononic systems. We obtain the entanglement entropy and entanglement spectrum for phononic systems with the fermion filling analog. With these measurements, we verify the Gioev-Klich-Widom scaling law. We further observe the salient signatures of topological phases in entanglement entropy and entanglement spectrum.

Observation of D-class topology in an acoustic metamaterial.

Topological materials and metamaterials opened new paradigms to create and manipulate phases of matter with unconventional properties. Topological D-class phases (TDPs) are archetypes of the ten-fold classification of topological phases with particle-hole symmetry. In two dimensions, TDPs support propagating topological edge modes that simulate the elusive Majorana elementary particles. Furthermore, a piercing of π-flux Dirac-solenoids in TDPs stabilizes localized Majorana excitations that can be braided for the purpose of topological quantum computation. Such two-dimensional (2D) TDPs have been a focus in the research frontier, but their experimental realizations are still under debate. Here, with a novel design scheme, we realize 2D TDPs in an acoustic crystal by synthesizing both the particle-hole and fermion-like time reversal symmetries for a wide range of frequencies. The design scheme leverages an enriched unit cell structure with real-valued couplings that emulate the targeted Hamiltonian of TDPs with complex hoppings: A technique that could unlock the realization of all topological classes with passive metamaterials. In our experiments, we realize a pair of TDPs with opposite Chern numbers in two independent sectors that are connected by an intrinsic fermion-like time-reversal symmetry built in the system. We measure the acoustic Majorana-like helical edge modes and visualize their robust topological transport, thus revealing the unprecedented D and DIII class topologies with direct evidence. Our study opens up a new pathway for the experimental realization of two fundamental classes of topological phases and may offer new insights in fundamental physics, materials science, and phononic information processing.

[Application of combined telomerase activity analysis and immunocytochemistry in cytopathologic diagnosis].

OBJECTIVE: To evaluate the application of traditional cytomorphology, telomerase activity analysis and immunocytochemistry in cytopathologic diagnosis of pleural effusion and bronchoalveolar lavage samples. METHODS: A total of 123 agar-paraffin double-embedded pleural effusion and bronchoalveolar lavage fluid samples were enrolled into study. The cytomorphologic features were reviewed and correlated with immunocytochemical findings and telomerase activity. RESULTS: Telomerase activity was detected in 53 specimens using the real-time telomeric repeat amplification protocol. Amongst the cases studied, 39 samples (31.7%) contained overtly malignant cells while 20 cases (16.0%) were equivocal by conventional cytology. After verification by immunocytochemistry and clinical follow-up data, the diagnostic accuracy of telomerase activity and cytology was 87.0% and 82.1%, respectively. The sensitivity (97.6%) and specificity (100.0%) of cytology examination, when combined with telomerase activity analysis, were greater than those of cytology examination or telomerase activity analysis alone. CONCLUSIONS: Telomerase activity analysis can be used as an adjunctive investigative tool in cytology assessment of pleural effusion and bronchoalveolar lavage samples. The diagnostic accuracy can be further improved with the application of immunocytochemistry on agar-paraffin double-embedded cell block tissues.

Detection of telomerase activity in cultured cells and tumor tissue of lung carcinoma by modified telomeric repeat amplification protocol.

Telomerase activity is found in various cell types including stem cells, neoplastic cells, and immortalized cells, suggesting a close association with their proliferation capacity. The telomeric repeat amplification protocol (TRAP) has been traditionally used to detect semi-quantitatively the telomerase activity by polyacrylamide gel electrophoresis (PAGE), which is difficult to apply for large scale analysis because of laborious post-PCR manipulation and potential carryover contamination. In the present study, a specific reverse primer was designed and the TRAP protocol was adapted to either PAGE or real-time PCR assay. Using cultured cell lines, the real-time TRAP showed a dramatic improvement in the reliability and accuracy of quantitation of telomerase activity and was able to discriminate the A549 cells from hundreds-fold human embryonic lung cells. Using clinical samples of 60 lung cancers and 8 inflammatory lesions, the real-time TRAP was also superior in quantitation, high-throughput capability and standardization. Our modified real-time TRAP should be applicable for the detection of telomerase activity for the initial screening and progression monitoring of lung cancer patients. Our approach is particularly useful when only limited clinical specimen is available, such as fine needle aspiration or other cytological specimens that may contain only a small number of tumor cells.

EGFR mutations are more frequent in well-differentiated than in poor-differentiated lung adenocarcinomas.

Somatic mutations in epidermal growth factor receptor (EGFR) tyrosine kinase domain, particularly deletions in exon 19 and point mutation in exon 21, are associated with clinical outcome in patients with lung adenocarcinoma, suggesting that EGFR mutation would have an important role in clinical decision making. DNA was extracted from the excised specimens of 60 lung adenocarcinoma patients with phenol-chloroform and ethanol precipitation. Exon 19 and 21 were amplified by PCR, and direct sequenced from both sense and antisense directions. EGFR somatic mutations were present in 13 of 60 patients (21.67%), including seven cases of in-frame deletion in exon 19 around codon 746 and six cases of amino acid substitution in exon 21. Exon 21 mutation is more frequent in adenocarcinomas with bronchi-alveolar component than exon 19 deletions. Mutations were more prevalent in well-differentiated adenocarcinomas (9/27, 33.33%) than in moderate to poor-differentiated adenocarcinomas (4/33, 12.12%) (P < 0.05). Adenocarcinomas with bronchi-alveolar components had higher mutation frequency (8/22,36. 36%) than those without bronchi-alveolar components (5/38, 13.16%) (P < 0.05). In this study, female patients had more mutation rate than male patients. This trend was also observed in the patients with pathologic stage I-II compared with stage III-IV, but neither of them was statistically significant. Patients with cisplatin-based adjuvant chemotherapy had no significantly prolonged survival compared with single radical resection. But patients with EGFR mutation had relative longer survival. In conclusion, our study suggest that EGFR mutations may be a valuable prognostic factor for disease free survival of surgically treated lung adenocarcinoma patients independently from adjuvant chemotherapy.

[Status and clinicopathologic implication of epidermal growth factor receptor mutation in non-small cell carcinoma of lung].

OBJECTIVE: To investigate mutations of epidermal growth factor receptor (EGFR) exon 19 and 21 in non-small cell lung carcinoma and to explore their clinicopathological correlations. METHOD: DNA was extracted from the excised tumor specimens of 66 non-small cell lung carcinoma patients by traditional phenol-chloroform and ethanol precipitation. Exons 19 and 21 were amplified by polymerase chain reaction (PCR), followed by direct sequencing in both sense and antisense directions. RESULTS: EGFR somatic mutations were present in 11 of 66 patients (16.7%), including 7 cases of in-frame deletion involving exon 19 and 4 cases of amino acid substitution involving exon 21. Mutations were more frequently observed in women (9/34, 26.5%) than in men (2/32, 6.3%), in adenocarcinomas (10/43, 23.3%) than squamous (0/13) and adenosquamous carcinomas (1/10). There was no difference in the mutation rates between smokers and non-smokers. Those with adenocarcinoma with bronchiolo-alveolar carcinoma (BAC) components had higher frequency of EGFR mutation (6/11) than those without non-BAC element (4/32, 12.5%). CONCLUSIONS: The mutations appear to occur in highly selected subgroups of lung cancer patients: adenocarcinomas with BAC components and patients of the female gender. The results may offer practical approach to the rapid identification of lung cancer patients who likely respond to EGFR inhibitor therapy.

[Detection of Mycobacterium tuberculosis DNA in sarcoidosis samples using real-time fluorescence polymerase chain reaction].

OBJECTIVE: To study the role of Mycobacterium tuberculosis in the pathogenesis of sarcoidosis. METHODS: Archival material of 22 patients with a histologic diagnosis of sarcoidosis were retrieved. Real-time fluorescent polymerase chain reaction (PCR) was used to detect DNA fragments of the complex-specific insertion sequence IS6110 of Mycobacterium tuberculosis in formalin-fixed and paraffin-embedded biopsy samples. RESULTS: Among the 22 samples studied, Mycobacterium tuberculosis DNA was detected in 11 cases. The sequence of PCR amplified IS6110 DNA fragments completely matched with the related sequence in Mycobacterium tuberculosis gene. CONCLUSIONS: Mycobacterium tuberculosis DNA is identified in a certain proportion of patients with a clinicopathologic diagnosis of sarcoidosis. Mycobacterium tuberculosis may be an important etiologic agent, at least in some of these patients.

[Status and clinical implication of c-kit and PDGFRA mutations in 165 cases of gastrointestinal stromal tumor (GIST)].

OBJECTIVE: To investigate the status of c-kit and PDGFRA mutations of GIST in a the large sample of Chinese patients. METHOD: One hundred and sixty-five cases were evaluated for the presence of c-kit and PDGFRA mutations. Exon 9, 11, 13, 17 of c-kit and exon 12, 18 of PDGFRA were analyzed by PCR amplification and direct sequencing. RESULTS: Immunohistochemical demonstrations of KIT (CD117) were seen in 94% of the cases (155/165). Overall, c-kit mutations were identified in 76.1% (118/155) of CD117 positive cases: 67.1% (104/155) involving exon 11, 7.1% (11/155) involving exon 9, 1.3% (2/155) involving exon 13 and 0.6% (1/155) involving exon 17. The c-kit exon 11 mutations were mostly heterogeneous and clustered in the classic "hot spot" at the 5' end of the exon, including in-frame deletion and point mutation. The second "hot spots" were internal tandem duplications (ITD) at the 3' end of the exon, which were associated with female patient, older age, stomach location and low mitotic counts. The exon 9 mutations correlated with a distinct subset of GISTs involving the small bowel of young male patients. A new point mutation of L641P was identified in exon 13. PDGFRA mutations were present in 50% (5/10) of CD117-negative GISTs, all involving exon 18 with the majority of mutations being D842V. One novel in-frame deletion of IMHD mutation at codon 843 - 846 with S847T was identified. GISTs with PDGFRA mutations were often larger tumors arising from the omentum/mesentery of young male patients with high risk of aggressive behavior. CONCLUSIONS: The vast majority of GISTs in this study harbored c-kit and PDGFRA mutations, there were non-random relations between the gene mutation patterns and the locations of GISTs. It appears that Chinese GIST patients have some unique mutation patterns. It is necessary to evaluate the gene mutations status of GISTs to guide target therapy.

[c-kit and PDGFRA mutations in 60 cases of gastrointestinal stromal tumors (GISTs)].

OBJECTIVE: To explore the status of activating mutations of c-kit and PDGFRA in GIST of Chinese patients. METHODS: Sixty GISTs, confirmed by immunoreactivity of CD117, CD34, SMA, S-100 and Desmin, were evaluated for the presence of c-kit exons 9, 11, 13 and 17 mutations, and PDGFRA exons 12 and 18 mutations. The PCR products were sequenced directly for mutations, using DNA extracted from paraffin-embedded tissue. RESULTS: 53% of the tumors were located in the stomach, 22% in the small bowel, 8% in the colo rectum, 2% in the esophagus and 15% in the extragastrointestinal tract. Immunohistochemical demonstrations of c-kit (CD117) were seen in 90% cases. Overall, c-kit mutations were detected in 63.3% of patients as follows: 58.3% in exon 11, 3.3% in exon 9, 1.7% in exon 13 and none in exon 17. The types of c-kit exon 11 mutations were mostly heterogeneous and clustered in the classic "hot spot" at the 5' end of exon 11, 42.9% being point mutation and in-frame deletion at Codon 557-560. 14.3% of cases showed internal tandem duplications (ITD) at the 3' end of exon 11 in a region of a second hot spot for c-kit mutations. Interestingly, these cases were associated with female predominance, stomach location and occurrence in older patients. The present study failed to identify a significant association between c-kit mutation status and risk of aggressive behavior in GISTs. Exon 9 mutations consisted of ITP of six nucleotides encoding Ala-Tyr. A new point mutation of L641P was revealed in exon 13. PDGFRA mutations were found in 5% of all the 60 cases with none of the positive cases expressed detectable KIT protein. The type of mutation was the commonest point mutation of D842V of exon 18. CONCLUSION: Most KIT expressing GIST show c-kit mutations that are preferentially located within the classic hot spot of exon 11. A second hot spot -ITD at the 3' end of exon 11 seems to associate with a subgroup of gastric GISTs in older females. c-kit exons 9, 13 and 17 mutations are rare events in GIST of China. PDGFRA oncogenic mutations are more likely seen in KIT-negative GISTs arising in the peritoneal surface and have an unfavorable clinical course.

Association of chemokine and chemokine receptor expression with the invasion and metastasis of lung carcinoma.

The chemokine system has been reported to be utilized and manipulated by tumor cells in order to promote local tumor growth and distant dissemination. The present study aimed to investigate the expression of three chemokine ligand-receptor axes in lung carcinoma tissues. Tumor and healthy normal tissue samples were obtained from 120 lung carcinoma patients following surgical resection. Immunohistochemistry and reverse transcription quantitative polymerase chain reaction were used in order to identify the protein and messenger (m)RNA expression of chemokines, including chemokine (C-X-C motif) ligand (CXCL)12/stromal cell-derived factor (SDF)-1, CXCL8/interleukin (IL)-8, chemokine (C-C motif) ligand (CCL)19 and CCL21, and the corresponding chemokine receptors, chemokine (C-X-C motif) receptor (CXCR)4, CXCR1, CXCR2 and chemokine (C-C motif) receptor (CCR)7, respectively. The results revealed that compared with the normal lung tissues, lung carcinoma tissues expressed significantly higher mRNA levels of CXCL12/SDF-1, CXCR4, CXCL8/IL-8, CXCR2, CCL19 and CCR7 (P<0.01). In four histological subtypes, adenocarcinoma presented dominant expression of CXCR4, CXCR2, CXCL8/IL-8 and CCL19 (P<0.05). In addition, it was demonstrated that tumor staging was inversely correlated with chemokine receptor CCR7 and CXCR2 mRNA expression as well as positively correlated with CXCL12/SDF-1, CXCL8/IL-8 and CCL19 mRNA levels (P<0.05). Lymph node metastasis presented a positive correlation with CXCR4, CXCR2 and CXCL8/IL-8 expression and a negative correlation with CCL19 and CCR7 expression (P<0.05). Furthermore, vascular invasion was more prevalent in patients with higher expression levels of CXCR4, CCR7 or CCL19 (P<0.01). In conclusion, these data suggested that the ligand-receptor interaction of CXCL8-CXCR2, CXCL12-CXCR4 and CCL19-CCR7 may be involved in the tumorigenesis of lung carcinoma. Higher expression levels of chemokines and lower expression of chemokine receptors indicated poor tumor staging. The CXC chemokine receptors, CXCR4 and CXCR2, promoted lymphatic metastasis through the activation of their specific ligands, while CCL19 and its receptor CCR7 had an essential role in hematogenous metastasis of lung carcinoma.

Helicobacter pylori infection and gastric cancer: evidence from a retrospective cohort study and nested case-control study in China.

AIM: To explore the association between Helicobacter pylori (Hp) infection and risk of gastric cancer in China. METHODS: Utilizing gastroendoscopic biospsy tissue banks accumulated from 1980 to 1988 in Shandong, Zhejiang, and Jiangsu, where stomach cancer incidence was high, during stomach cancer screening conducted by Health Science Center of Peking University, School of Medicine of Zhejiang University, and Zhongshan Hospital of Fudan University. Warthin Starry silver staining method was applied to determine H. pylori infection status of biopsies collected during gastroendoscopic examination. In the retrospective study, the subjects were divided into two cohorts, the exposure cohort was positive H. pylori infection, and the non-exposure cohort was negative. Death from stomach cancer was determined as the outcome of the study. Logistic regression and Cox regression were applied to analyze the association between Helicobacter pylori infection and gastric cancer risk. In the nested case-control study, there were 28 deaths from gastric cancer in the fields of Muping, Shandong province, and Zhoushan, Zhejiang provinces. 4 controls were matched to each case on the basis of age (+/-5 years old), sex, residential place at the same time entered into the study. Conditional logistic regression analysis was used to analyze the data. RESULTS: There were a total of 2 719 subjects (male 1 399, female 1 320) with gastroendoscopic biopsies stored available treated as a cohort. H. pylori positive cohort included 1 671 subjects (61.5 %) and H. pylori negative cohort 1 048 subjects(38.5 %). These subjects were followed up for 1-19 years, averaged 10.88 years. The outcome of death from stomach cancer in the exposure cohort was 33, and in the non-exposure cohort 11. After adjustment for age and sex, RR=1.9850 (P=0.0491), 95 % CI was 1.0026, and 3.9301. The results of conditional logistic regression showed an OR of 4.467 and 95 % CI of 1.161, and 17.190 for the nested case control study. CONCLUSION: The results from the retrospective cohort study and the nested-case control study on the association of H. pylori infection and gastric cancer in China suggested that Helicobacter pylori infection might increase the risk of stomach cancer.

[Effects of two variants of ING1 expression on tumor cell growth regulation].

OBJECTIVE: To study effects of alternative transcripts of ING1 transfection on human cancer cell lines. METHODS: p47/ING1A and p33/ING1B expression vehicles were constructed and introduced into a human breast cancer cell line MCF-7 and a human lung cancer cell line PAa, both expressing wild-type p53 protein. Growth characteristics of the transfectants and potentially related genes were analyzed. RESULTS: The levels of p47/ING1A and p33/ING1B protein elevated respectively in tumor cells of MCF-7 and PAa after transfected with p47/ING1A and p33/ING1B, and the latter was much higher than that of the former. Ectopic overexpression of p33/ING1B effectively blocked tumor cell growth and arrested cells in the G(0) approximately G(1) phase of the cell cycle (P < 0.01), while p47/ING1A gave no effect on cell growth or cell cycle. Tumor cells overexpressing p33/ING1B contained more p21(WAF1) protein than that of the control cells, with undisturbed p53 protein level. CONCLUSIONS: Expression of two different transcripts of ING1 may have different effects on tumor cell growth. p33/ING1B may cooperate with p53 in stimulating expression of p21(WAF1) gene, thus to arrest cell cycle and to inhibit tumor cell growth. p33/ING1B may be considered to be a candidate as a partner of p53 in gene therapy.

[Gene detection of severe acute respiratory syndrome-related coronavirus].

OBJECTIVE: To develop a newly real-time RT-polymerase chain reaction assay for severe acute respiratory syndrome (SARS) related coronavirus in human whole blood. METHODS: A pair of primers and a probe (molecular beacon) had been designed that were specific for the recognition of a highly conservative region between 15 301 and 15 480 of the SARS-related coronavirus polymerase gene sequences obtained from GenBank (G130027616). RESULTS: In the real-time RT-PCR assay, the extent of SARS related coronavirus amplification was measured in terms of the increase in fluorescence during the amplification process. The 145 bp fragment of PCR product was further confirmed by conventional PCR assay and proved by DNA sequencing to be identical to the target sequence to which the probe was hybridized. CONCLUSION: This assay has a broad application for clinical diagnosis and surveillance investigation.

[A novel Krit-1 mutation in Han family with cerebral cavernous malformation].

OBJECTIVE: To detect the mutations of Krit-1 gene that cause familial cerebral cavernous malformation (CCM) in the Han ethnic origin. METHODS: The subjects were hospitalized in the Department of Neurosurgery, Tiantan Hospital affiliated to Capital University of Medical Sciences. Two families (A and B) and 8 apparently sporadic individuals affected with CCM were screened for mutations of Krit-1 gene. Members of the family CCM have a wide range in age of onset with seizures, headaches and skin lesions. The gene was screened by PCR amplification of 16 exons and mutation was detected by direct sequencing. RESULTS: In family A samples, analysis of the Krit-1 gene revealed a new point mutation in exon 14 [a heterozygous C to G transition at nucleotide 1 289 (counting from the start codon or nt 2 308 counting from the first nt of the mRNA, aligned according to Gene Bank AF388384)] which predicts the substitution of a premature termination codon for Serine at codon 430 (S430X), belonging a nonsense point mutation. No mutation was identified in one of family A members as well as in any of the sporadic individuals with the exception of a single nucleotide polymorphism. CONCLUSIONS: Report the first family in the Han with CCM having a novel mutation in the CCM1 gene on the continent of Asia. The newly identified mutation creates a premature termination codon and is predicted to produce a truncated Krev1 interaction-trapped 1 protein, KRIT1. This result allows efficient presymptomatic molecular diagnosis.

STR DNA genotyping of hydatidiform moles in South China.

OBJECTIVE: To evacuate whether short-tandem-repeat (STR) DNA genotyping is effective for diagnostic measure to precisely classify hydatidiform moles. METHODS: 150 cases were selected based on histologic features that were previously diagnosed or suspected molar pregnancy. All sections were stained with hematoxylin as a quality control method, and guided the microscopic dissection. DNA was extracted from dissected chorionic villi and paired maternal endometrial FFPE tissue sections. Then, STR DNA genotyping was performed by AmpFlSTR(®) Sinofiler(TM) PCR Amplification system (Applied Biosystems, Inc). Data collection and analysis were carried out using GeneMapper(®) ID-X version 1.2 (Applied Biosystems, Inc). RESULTS: DNA genotyping was informative in all cases, leading to identification of 129 cases with abnormal genotype, including 95 complete and 34 partial moles, except 4 cases failed in PCR. Among 95 complete moles, 92 cases were monospermic and three were dispermic. Among 34 partial moles, 32 were dispermic and 2 were monospermic. The remaining 17 cases were balanced biallelic gestations. CONCLUSION: STR DNA genotyping is effective for diagnostic measure to precisely classify hydatidiform moles. And in the absence of laser capture microdissection (LCM), hematoxylin staining plus manual dissection under microscopic guided is a more economic and practical method.