Advances in Clinical Application and Hemostatic Mechanism of Charcoal Drugs of Chinese Materia Medica / 中国实验方剂学杂志

Under the guidance of the theory of traditional Chinese medicine （TCM）， charcoal drugs are widely used in clinical treatment of various bleeding syndromes， in addition， they also have the effect in anti-diarrhea and anti-ulcer， but charcoal drugs are especially effective in stopping bleeding. According to the changes in the properties after processing， the hemostatic effect of charcoal drugs can be roughly divided into two categories. One is not used for hemostasis itself， but used for hemostasis after processing. The other is used for hemostasis itself， and the drug properties are changed or the hemostatic ability is enhanced after processing. By summarizing researches on historical evolution， processing mechanism and pharmacological effects of the commonly used hemostatic charcoal drugs， the author found that preservation or increase of active substances after processing was closely related to the hemostatic effect of charcoal drugs. The hemostatic mechanism mainly involves the influence of coagulation system and platelet function， etc. At the same time， combined with the theory of Qi chromatograph of TCM supramolecular， this paper puts forward the supramolecular research strategy on hemostatic mechanism of charcoal drugs， in order to provide reference for revealing the scientific connotation of charcoal drugs for hemostasis.

Construction of Evaluating Method and Experimental Research for Targeting of Traditional Chinese Medicine Preparations Based on Supramolecular Qi Chromatography Theory / 中国实验方剂学杂志

Objective:To construct the targeting evaluation method of traditional Chinese medicine （TCM） preparations based on supramolecular Qi chromatography theory， and to study the liver targeting effect of Bupleuri Radix on Pien Tze Huang. Method:The molecular connectivity index （MCI） was used to analyze the characteristics of imprinted template and liver targeting tendency of TCM mainly attributed to liver meridian and components of Pien Tze Huang， and combined with target dynamics and total statistical moment principle， aimed at the independent action characteristics of multi-component imprinted template of TCM， a method for evaluating the targeting of TCM preparations was established. Hepatoma rats in Pien Tze Huang group， Bupleuri Radix<italic> </italic>group， Pien Tze Huang+Bupleuri Radix group and blank group were tested and verified. Result:After the average value of MCI of TCM mainly attributed to liver meridian was deducted， the MCI similarity between Pien Tze Huang group and Bupleuri Radix group was 0.376 8， Pien Tze Huang+Bupleuri Radix group and Bupleuri Radix group was 0.988 2， so it was predicted that Bupleuri Radix could enhance the liver targeting of Pien Tze Huang. A system for evaluating the targeting of TCM compounds was established， including relative total uptake efficiency （RUE_T）， relative total concentration （RC_T）， relative imprinted tendency （RIT_T） and relative imprinted variance （RIV_T）. The RUE_T and RC_T of liver were the highest in all tissues （RUE_T=1.88>1，RC_T<italic>=</italic>2.30>1）， and the corresponding values of other tissues were all <1， indicating that Pien Tze Huang combined with Bupleuri Radix could increase its distribution in liver and enhance liver targeting. Except for plasma， the RIT_T and RIV_T of other tissues fluctuated around 1.0， indicating that targeted modification did not change imprinted tendency of Pien Tze Huang and had no significant effect on the types of components. Conclusion:Under the guidance of supramolecular Qi chromatography theory， a targeting evaluation parameter system can be established to characterize the multi-component imprinted effect of TCM preparations by MCI and total statistical moment parameters， so as to realize the evaluation of targeting of TCM preparations. The addition of Bupleuri Radix can increase the liver targeting of Pien Tze Huang.

Research status and regulatory mechanism based on supramolecular "imprinted template" of toxicity of Chinese materia medica / 中草药

Based on the universal regularity of the automatic action of supramolecular "imprinted template" previously proposed, this paper further analyzed and summarized the historical evolution and research status of the toxicity of Chinese materia medica, and deeply analyzed the production of toxicity of Chinese materia medica by using the theory of supramolecular chemistry. After entering the human body, the aggregation of supramolecular "imprinted template" of Chinese materia medica can interact with the similar "imprinted template" of the subject supramolecular of human in the human body. In other words, the group of toxic components of Chinese materia medica can be identified with the corresponding hole channels in the body, which is the supramolecular chemical mechanism of the toxicity of Chinese materia medica. The dose-effect problem and detoxification treatment of toxic Chinese materia medica is closely related to the equilibrium constant and concentration. When the concentration of the drug with the same (similar to) imprinted template is the same, the toxicity depends on the equilibrium constant of its interaction with the target. Based on this, research idea of equilibrium constant of toxicity of Chinese materia medica was proposed. The equilibrium constant of supramolecular imprinted behavior can explain the toxicity of Chinese materia medica. By studying the equilibrium constant, the supramolecular interaction of the toxic effect can be revealed and the supramolecular system of Chinese materia medica can be well studied.

Investigation of Role of TCM Supramolecular "Qi Chromatography" in Modernization of TCM Theory of Epidemic Diseases Caused by SARS-CoV-2 / 中国实验方剂学杂志

In this paper, based on the supramolecular "Qi chromatography" of traditional Chinese medicine (TCM), combined with the characteristics of corona virus disease 2019 (COVID-19), the historical origin and characteristics of theoretical research on the treatment of epidemic diseases by TCM were analyzed retrospectively, and the key scientific problems for achieving its modernization were proposed, and the countermeasures were illustrated. In fact, the emergence and development history of TCM epidemic theory is a comprehensive and scientific understanding of biomolecular chemistry for epidemic caused by pathogen. From the exploration of the cause of abnormal Qi to the epidemic pathogenic factor, from the treatment based on syndrome differentiation by six meridians to by the defensive Qi and nutrient blood and the triple energizer, which is gradually close to the modern theoretical system of infectious diseases and immunology, all of which are stamped with brand of TCM supramolecular "Qi chromatography" theory. In the long history of biological evolution in nature, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) as epidemic pathogenic factor in TCM, human body and Chinese materia medica are biological supramolecules at a specific stage of evolution in nature, which are controlled by the "imprinting templates" of molecular society in gradual biomolecules, with the unchanged laws of mutual advancement, mutation, suitability and mutual restriction. Based on the analysis of the law of supramolecular action hiding in the prevention and treatment of epidemic diseases dealt with TCM on procession of principle-method-recipe-medicines, this paper discusses the key scientific problems of its modernization and puts forward the solutions. It can not only clarify the scientific principle of the prevention and treatment with TCM, develop the multi-component Chinese medicine, accelerate the control of the epidemic situation, but also promote the solution of original frontier key scientific problems such as the material basis of meridians, viscera and syndromes, and the compatibility law of the TCM prescription, in order to protect the existing medical resources and open up a new situation in the modernization of TCM research.

Quality Control of Dynamic Volatile Components in Lonicerae Japonicae Flos and Lonicerae Flos / 中国实验方剂学杂志

Objective:The molecular connectivity index method and total statistical moment method were used to control the quality of traditional Chinese medicine （TCM）， and the stability and consistency of volatile components of Lonicerae Japonicae Flos and Lonicerae Flos were clarified. Method:Volatile oils in Lonicerae Japonicae Flos and Lonicerae Flos from different producing areas was extracted for GC-MS determination with electron bombardment ion source， ion source temperature of 230 ℃， and detection range of m/z 35-650. Then National Institute of Standards and Technology （NIST） 05 and ChemicalBook database were used for qualitative analysis of these volatile components， the peak area normalization method was used for quantitative analysis， and the total statistical moment parameters and the zero-order， first-order， second-order， third-order molecular connectivity indexes of the components were calculated. Result:Number of peaks （RSD were 28.5%， 33.4%， respectively）， total zero-order moments （RSD were 55.5%， 128.9%， respectively） and total second-order moments （RSD were 15.3%， 21.5%， respectively） of 10 batches of Lonicerae Japonicae Flos and Lonicerae Flos were unstable， indicating that the types and contents of volatile components fluctuated sharply， but the total first-order moments （RSD were 7.5%， 8.8%， respectively） and the zero-order， first-order， second-order and third-order molecular connectivity indexes （RSD ranged from 8.1% to 10.3% and 4.2% to 5.5%， respectively） were relatively stable， indicating that the overall "imprinting template" of the components was similar. Statistical analysis of each parameter found that there were no significant differences in the number of peaks， total first-order moments and zero-order， first-order， second-order， third-order molecular connectivity indexes between volatile oils from Lonicerae Japonicae Flos and Lonicerae Flos. Conclusion:Under the guidance of supramolecular gas evolution "imprinting template" theory， the molecular connectivity index method and total statistical moment parameters are used to jointly characterize the "imprinting template" of TCM components in vitro， which can control the stability and consistency of TCM quality.

Analysis of Key Techniques in Development of Primary Standard of Classical Prescription of Chinese Medicine / 中国实验方剂学杂志

Research and development of classical prescription of Chinese medicine has become one of the hot spots in the research of traditional Chinese medicine (TCM), and the successful development of the primary standard is crucial to the application of the classical prescription of Chinese medicine. Primary standard of classical prescription is not only the benchmark to measure the quality of preparation of classical prescription, but also the material basis of whole prescription. The development and quality control mode of single component of western medicine is not suitable for the development of TCM preparation with integral medicinal function. Thus, it is very urgent to develop an exclusive research and development model for TCM. In order to break through the quality control technology of primary standard of classical prescription, and comprehensively and accurately elucidate the transmission rule of the component group value of medicinal materials-decoction pieces-primary standard of classical prescription with the present modern science and technology, the genetic polymorphism of TCM, extraction kinetics, total quantum statistical moment (similarity) method of fingerprint and supramolecular imprinting template were combined and applied to the development of primary standard of classical prescription. In this way, the development and quality control of TCM will be realized in a real sense and the development of classical prescription of Chinese medicine will be accelerated and promoted.

Analysis of Total Statistical Moment and One-time Steady-state Feeding Quantity of HPLC Fingerprint of Taohong Siwutang / 中国实验方剂学杂志

Objective:To establish a mathematical model of total statistical moment, information amount and one-time steady-state feeding quantity of HPLC fingerprint of traditional Chinese medicines (TCMs) and their classical prescriptions, and to carry out experimental verification. Method:The total statistical moment principle, Hardy-Weinberg equilibrium principle and one-time steady-state feeding quantity calculation method were used to study the total statistical moment parameters and one-time steady-state feeding quantity of 15 batches of original medicinal materials, decoction pieces and compound preparations of Taohong Siwutang, these original medicinal materials were from three producing areas. Result:In addition to the total first-order moment, RSD of total zero-order moment, total second-order moment, information entropy and information amount of each medicinal material, decoction piece and compound were all greater than 10%. In the case of single original medicinal material, except for Angelicae Sinensis Radix (142.34 kg), the one-time steady-state feeding quantity calculated according to the RSD change of the accumulated information amount of the whole prescription was the largest (59.10 kg), and according to ratio of the whole prescription, one-time steady-state feeding quantity was the largest (958.57 kg). In the case of processed products, the one-time steady-state feeding quantity of the whole prescription was the largest (73.18 kg) except for Carthami Flos processed with wine (83.28 kg), while according to the whole prescription, one-time steady-state feeding quantity of each processed products was the same as that of the original medicinal material. The one-time steady-state feeding quantity of the compound consisted of the original medicinal materials or processed products was often higher than that of the single medicinal material. Conclusion:The components of TCM are controlled by genetic polymorphism, the chromatographic characteristics (types of components) of Chinese medicine is similar and the intensity of action (content of components) is different. The stability of total first-order moment and information entropy indicates that the overall chemical structure of TCM multi-component system has a stable imprinting trend. The one-time steady-state feeding quantities of each medicinal material in Taohong Siwutang and this compound are far greater than the prescription amount, which can provide reference for the industrial production.

Research on Quality Control Model of Ancient Famous Classical Formulas Based on Supramolecular Theory / 中国实验方剂学杂志

The ancient famous classical formulas were developed based on the characteristics of excellent curative effect, small toxic and side effects, and wide clinical application. Its development will promote the development of traditional Chinese medicine (TCM) with huge social and economic benefits. TCM and TCM compounds are multi-component systems, which are multi-component, multi-target and multi-pathway effect on human body, their curative effect is the result of synergistic effect of multi-component. Because of its characteristics, its mechanism of action is still unclear and quality is difficult to control. Therefore, on the basis of biological properties of the multi-components of TCM, it is the key issue to study the preparation process of the medicinal materials-decoction pieces-preparations and the transferability, stability and homogeneity of the quality attributes. Moreover, the supramolecular theory can explain the basic theory of TCM from the macroscopic and microscopic aspects, the study of ancient famous classical formulas can use the supramolecular theory. Both TCM and human body are giant complex supramolecules, and their components are clustered objects reflecting the characteristics of the original imprinting template with genetic polymorphism. The quality attributes, processing, formulation preparation and quality attribute evaluation of TCM are all affected by supramolecular chemistry, which is finally reflected in the preparation of imprinting template and the transfer law of quality properties. Therefore, the application of supramolecular imprinting template theory to study the biogenetic polymorphism of TCM, processing, formulation preparation and quality attribute evaluation of medicinal materials can improve the availability rate of successful research and development of ancient famous classical formulas.

Photodynamic Therapy Mediated by Aloe-Emodin Inhibited Angiogenesis and Cell Metastasis Through Activating MAPK Signaling Pathway on HUVECs.

Photodynamic therapy is a clinically used, minimally invasive therapeutic procedure that involves the application of photosensitizers which can locate in target cells and so be irradiated at a corresponding wavelength. Laser light irradiation activation of photosensitizers generates free reactive oxygen species, which induces selective cytotoxic activity in target cells. Within recent years, aloe-emodin as a photosensitizer has been successfully applied in photodynamic therapy applications. Angiogenesis plays an important role in tumor growth and metastasis; thus, the development of a novel target treatment for angiogenesis is essential in order to improve treatment therapeutics for cancer treatment. An essential step in angiogenesis involves the formation of tube-like structures during matrix degradation, rearrangement, and apoptosis of endothelial cells. In the present study, we investigated the mechanisms of photocytotoxicity induced by aloe-emodin in human umbilical vein endothelial cells. Analysis of cell proliferation results noted a significant decrease in cultured cells which received various concentrations of aloe-emodin and photodynamic therapy-induced light doses. Additionally, mitochondrial mechanisms of apoptotic cell death were observed in aloe-emodin photodynamic therapy-treated cells, as tube formation assays noted angiogenesis suppression after treatment. The capacity of migration and invasion of human umbilical vein endothelial cells was measured using the transwell assay and demonstrated that aloe-emodin photodynamic therapy significantly inhibited the migration and invasion of human umbilical vein endothelial cells. The expression of p38, extracellular signal-regulated kinase, the c-Jun N-terminal kinases, and vascular endothelial growth factor suggested that the cellular metastasis was related to mitogen-activated protein kinase signal pathway. Furthermore, disorganization of F action cytoskeleton components was observed after aloe-emodin photodynamic therapy. Overall, the findings from this study suggest that aloe-emodin photodynamic therapy inhibited angiogenesis and cellular metastasis in human umbilical vein endothelial cells by activating the mitogen-activated protein kinase apoptotic signaling cell death pathway.

Enhancement of the Effect of Methyl Pyropheophorbide-a-Mediated Photodynamic Therapy was Achieved by Increasing ROS through Inhibition of Nrf2-HO-1 or Nrf2-ABCG2 Signaling.

BACKGROUND: Emerging evidence indicates that the transcription factor nuclear factor-E2-related factor 2 (Nrf2) plays an essential role in cellular defense against oxidative stress; its activation has been related to cytoprotection. OBJECTIVE: Here, we investigated the role of Nrf2 in improving the efficacy of methyl pyropheophorbide-amediated photodynamic therapy (Mppa-PDT) via the downregulation of Nrf2. METHOD: Human ovarian cancer A2780 cells and SKOV3 cells were treated with Mppa-PDT and siRNA transfection was performed to inhibit Nrf2. After treated with siRNA and Mppa-PDT, the cell viability was examined with CCK-8 assay; cell apoptosis was detected tested by flow cytometry with Annexin V-FITC/PI; the celluar reactive oxygen species (ROS) and mitochondrial membrane potential were measured with DCFHDA and JC-1 staining; expression of protein was assessed by western blot analysis. RESULTS: We found that Nrf2 translocated from the cytoplasm to the nucleus in vitro and in vivo, and the expression of Nrf2 and P-Nrf2 increased through a possible mechanism regulated by mitogen-activated protein kinase (MAPK) after Mppa-PDT treatment. Furthermore, cytotoxicity and apoptosis induced by Mppa-PDT increased after Nrf2down-regulation. Nrf2 down -regulation increased reactive oxygen species (ROS) levels by attenuating antioxidants or pumping Mppa out of cells,which resulted from the inhibition of Nrf2-HO-1 or Nrf2- ABCG2 signaling. In addition, SKOV3 cells exhibited increased resistance to Mppa-PDT, and the expression levels of P-Nrf2 and ABCG2 were higher in SKOV3 cells than in A2780 cells, suggesting that Nrf2-ABCG2 signaling might be involved in the intrinsic resistanceto Mppa-PDT. CONCLUSION: These results provided evidence that Nrf2 down-regulation can enhance the effect of Mppa-PDT.

[Mitochondria-targeted antioxidant Mitoquinone protects post-thaw human sperm against oxidative stress injury].

OBJECTIVE: To investigate the potential protective effect of the mitochondria-targeted antioxidant Mitoquinone (MitoQ) on post-thaw human sperm. METHODS: Semen samples were collected from 60 normal fertile men, each divided into six parts of equal volume to be incubated at 37 °C in normal saline (G0, control) or in the extender with 2 nmol/L (G1), 20 nmol/L (G2), 200 nmol/L (G3), 2 µmol/L (G4), and 20 µmol/L of MitoQ (G5). After one hour of incubation, the samples were subjected to computer-assisted semen analysis (CASA) for sperm motility, flow cytometry for reactive oxygen species (ROS), thiobarbituric acid assay for the concentration of malondialdehyde (MDA), and MitoTracker fluorescent staining and flow cytometry for the sperm mitochondrial membrane potential (MMP). Then, the semen were cryopreserved with none (B0), 200 nmol/L (B1), and 2 µmol/L of MitoQ (B2), followed by detection of the changes in the ROS, MDA, and MMP of the post-thaw sperm. RESULTS: The percentage of progressively motile sperm and total rate of sperm motility were significantly higher in G3 ([30.8 ± 10.2]% and [70.6 ± 9.0]%) and G4 ([32.7 ± 13.5]% and [70.3 ± 11.9]%) than in G0 ([17.6 ± 5.0]% and [54.9 ± 11.5]%) (P < 0.05). The level of ROS dropped markedly with the increased concentration of MitoQ, 86.5 ± 31.6 in G3, 93.6 ± 42.0 in G4, and 45.1 ± 15.0 in G5, as compared with 160.8 ± 39.7 in G0 (P < 0.05). The content of MDA was remarkably lower in G3 ([0.9 ± 0.5] µmol/mg) and G4 ([0.9 ± 0.5] µmol/mg) than in G0 ([1.9 ± 1.1] µmol/mg) (P < 0.05), but not in G5 ([1.7 ± 0.7] µmol/mg), which was even higher than in G3 and G4 (P < 0.05). The MMP showed a significant reduction in G5 (1156 ± 216) in comparison with G0 (1701 ± 251) (P < 0.05) but exhibited no remarkable difference between G0 and G1 (1810 ± 298), G2 (1995 ± 437), G3 (1950 ± 334), or G4 (1582 ± 314). The percentage of progressively motile sperm and total rate of sperm motility after freezing-thawing were significantly decreased as compared with those of the fresh semen (P < 0.01), but both were remarkably higher in B1 ([3.2 ± 2.3]% and [ 43.0 ± 9.5]%) than in B0 ([0.8 ± 0.6]% and [26.5 ± 11.4]%) (P < 0.05). The ROS level was significantly lower in B1 and B2 than in B0 (34.6 ± 12. 3 and 37.0 ± 10.5 vs 56.9 ± 14.3, P < 0.05), and so was the MDA content ([1.4 ± 0.5] and [1.4 ± 0.6] µmol/mg vs [2.6 ± 1.0] µmol/mg, P < 0.05), but the MMP was markedly higher in B1 and B2 than in B0 (1010.0 ± 130.5 and 880.6 ± 128.6 vs 721.1 ± 24.8, P < 0.05). CONCLUSION: Addition of MitoQ to the freezing extender at 200 nmol/L may effectively improve the quality of human sperm and MitoQ is a good protective addictive for human sperm cryopreservation.

Exploring a Novel Target Treatment on Breast Cancer: Aloe-emodin Mediated Photodynamic Therapy Induced Cell Apoptosis and Inhibited Cell Metastasis.

Photodynamic therapy (PDT) as a clinical cancer therapy, is a mild therapy, which involves application of photosensitizers (PSs) located in target cells and then irradiated by corresponding wavelength. The activation of PSs generates radical oxygen species (ROS) to exert a selective cytotoxic activity for the target cells. Aloe-emodin (AE) has been found to be an anti-tumor agent in many studies, and has also been demonstrated as a photosensitizer, in the recent years. In order to study the mechanisms of aloe-emodin as a photosensitizer, we investigated the mechanisms of photo-cytotoxicity induced by aloe-emodin in breast cancer MCF-7 cells in the present study. Analysis of cell proliferation evidenced that there was a drastic depression after photodynamic treatment with a series of aloe-emodin concentrations and light doses. We observed changes in apoptosis and demonstrated that the mechanisms of apoptosis were involved in mitochondrial and endoplasmic reticulum death pathways. The capacity of adhesion, migration and invasion of breast cells was measured using WST8 and transwell assay and demonstrated that AE-PDT significantly inhibited adhesion, migration and invasion of MCF-7cells. The expression of MMP2, MMP9, VEGF and Nrf2 demonstrated that the metastasis was related to oxidative stress. Analysis of changes in cytoskeleton components (F-actin) evidenced cytoskeleton disorganization after treatment with AE-PDT. Taken together, the present results indicated that PDT with aloe-emodin effectively suppressed cancer development in MCF-7cells, suggesting the potential of AE as a new photosensitizer in PDT which can provide a new modility for treating cancer.

Antigenic comparative analysis of Newcastle disease viruses with evolutional mutations in HN and F genes under antibody immune pressures / 病毒学报

In chicken fibroblast cell (CEF) cultures with antiserum against Newcastle disease virus (NDV) strain TZ060107, the virus was passed serially for 50 passages in 3 independent lineages. HN and F genes were amplified and sequenced every 10 passages. The derived virus A1-50 with most mutations among 3 lineages was further passed for another 50 passages in CEF with or without antiserum against A1-50, each in 3 independent lineages. Sequence comparisons for HN and F genes of 60, 70, 80, 90 and 100 passages indicated that the ratio of nonsynonymous mutations (NS) vs synonymous mutations (S) for HN genes in the lineages passed with antiserum against A1-50 was 5.25, which was obviously higher than 2. 375 of NS/ S in the lineages without the antiserum. The stable NS mutations occurred in the first 50 passages with the antiserum against the original TZ060107 were still maintained and one more new stable NS mutation appeared. For the F gene, 3 new stable NS mutations occurred during the second 50 passages in lineages with antiserum against A1-50 when the original NS mutations obtained in the first 50 passages with antiserum against TZ060107 still existed. Cross hemagglutination inhibition (HI) between original virus and its derivative viruses indicated that the more continuous passages in cell culture with antiserum passed, the bigger difference of antigenicity between the virus and the original virus had.

High expressions of bcl-2 and survivin, and decreased apoptosis in uterine cervical carcinosarcoma compared to cervical squamous cell carcinoma.

PURPOSES: Uterine cervical carcinosarcoma (CS) is very rare. To date, only 40 cases have been reported. It seems to have a more aggressive clinical behavior than does cervical squamous cell carcinoma (SCC). The purposes of our study were to characterize the clinicopathologic characteristics and human papillomavirus (HPV) status of the rare tumor and to analyze the molecular features in cervical CS that may account for its aggressive behavior. METHODS: Three patients were diagnosed with uterine cervical CS at West China Second Hospital of Sichuan University between 1995 and 2009. Data were retrospectively analyzed from available charts and pathological reports. Twelve patients with FIGO stage Ib-IIa cervical SCC were enrolled as the controls, and the expression profiling of p53, Ki-67, bcl-2, survivin and apoptosis index between cervical CS and SCC was compared. Immunohistochemical and apoptosis results were scored separately for the carcinomatous and sarcomatous components. RESULTS: All three patients were shown to be negative for HPV infection by Hybribio HPV genoarray assay. Expression of p53 was observed in one patient in both carcinomatous and sarcomatous components in a similar proportion; in contrast, the Ki67, bcl-2 and survivin expressions were higher in carcinomatous components than in sarcomatous components in all three cases. Compared to cervical SCC, stronger immunostaining for bcl-2, survivin and lower apoptosis was observed in cervical CS. CONCLUSIONS: Cervical CS is a peculiar tumor with many different clinicopathologic characteristics from cervical SCC. Dysregulation of apoptosis may confer tumor cells of cervical CS with survival and growth advantages, and thereby facilitate the aggressive behavior of cervical CS.

Association of an interleukin-16 gene polymorphism with the risk and pain phenotype of endometriosis.

Interleukin-16 (IL-16), a proinflammatory cytokine, plays a pivotal role in inflammatory diseases as well as in the pathogenesis of endometriosis. The aim of this study was to evaluate the association of IL-16 gene polymorphisms with the risk and clinical phenotypes of endometriosis in Chinese women. We analyzed rs4778889 T/C, rs11556218 T/G polymorphisms of the IL-16 gene in 230 patients with endometriosis and 203 controls in a Chinese population, using a polymerase chain reaction-high resolution melting analysis strategy and DNA sequencing methods. There was no significant difference in the genotype and allele frequencies of the rs11556218 T/G polymorphism between patients with endometriosis and controls (p>0.05). In contrast, the genotype and allele frequencies of the rs4778889 T/C polymorphism were statistically different between patients with endometriosis and controls, which resulted from a significantly increased proportion of TC heterozygote and CC homozygote carriers among patients with endometriosis (p=0.001 and 0.012, respectively); moreover, further subgroup analysis found that the genotype difference was more evident in patients with endometriosis who also experienced pain symptoms (p<0.001) than in patients without pain symptoms (p=0.625) when compared with controls. Our results suggest that the rs4778889 T/C polymorphism of the IL-16 gene may be associated with risk of endometriosis in the Chinese population, especially in patients with pain phenotype.

Immunological properties of recombinant Mycobacterium bovis bacillus Calmette-Guérin strain expressing fusion protein IL-2-ESAT-6.

The live vaccine Mycobacterium bovis bacillus Calmette-Guérin (BCG) provides variable efficacy against adult pulmonary tuberculosis (TB). Recombinant BCG, expressing either immunodominant antigens or Th1 cytokines, is a promising strategy for developing a new TB vaccine. However, not much is known about whether the introduction of cytokine and specific antigen genes concurrently into the BCG strain could improve the immunogenicity of BCG. In this study, a recombinant BCG strain (rBCG) expressing the fusion protein human interleukin (IL)-2 and ESAT-6 (early secreted antigenic target-6 kDa) antigen of Mycobacterium tuberculosis was constructed. Six weeks after BALB/c mice (H-2d) were immunized with 106 colony forming units (CFUs) BCG or rBCG, splenocyte proliferation was determined with MTT [3-(4,5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide] assay, IL-4 and interferon (IFN)-gamma produced by splenocytes were tested by enzyme linked immunosorbent assay (ELISA,) and the cytotoxicity of splenocytes from immunized mice to P815 cells (H-2d) expressing ESAT-6 protein was measured using CytoTox 96 Non-Radioactive Cytotoxicity Assay. Compared with native BCG-vaccinated mice, rBCG induced stronger Th1 responses that were confirmed by high lymphoproliferative responses and IFN-gamma production to culture filtrate protein (CFP) or ESAT-6 protein. Moreover, rBCG induced significant enhanced CTL responses against P815-ESAT-6 cells. Results from rBCG-immunized mice demonstrated that introducing the il-2 and esat-6 genes into BCG could enhance Th1 type immune responses to ESAT-6. Further investigation is needed by introducing other Th1 cytokines and antigens into BCG to optimize the protective efficacy against TB.