Two active compounds were isolated from chinese marine mollusc sea hare (Notarchus leachii cirrosus) by bioassay guided method with conidia of Pyricularia oryzae , and by a combination of extraction and purification with suitable solvent as well as multiple column chromatographies. Their structures were elucidated as (7 E) 1 acetoxy 8 chloro 7 (dichloromethyl) 3 methyoct 7 en 4 one (Ⅰ), (7 Z) 1 acetoxy 8 chloro 7 (dichloromethy l) 3 methyoct 7 en 4 one (Ⅱ). All compounds were obtained for the first time from this specimen and showed significant antineoplastic activities in vitro .

Objective:To screen for the bioactive marine organisms and to study the active constituents from them.Methods:Bioactivities of marine organisms extracts were screened by Pyricularia oryzae bioassay method and the constituents were isolated and identified by a combination of multi chromatography and spectral analysis.Results:Twenty ethanolic extracts and 5 aqueous extracts of the tested marine organisms showed activities causing morphological abnormality of P.oryzae mycelia.Fifty six compounds were obtained and identified from Sargassum carpophyllum,Ishige okamurai and Natarchus leachii freeri ,among which 28 compounds showed activities against P.oryzae and exhibited cytotoxicities on various cultured tumor cell lines,eight compounds showed inhibition effects on Fonsecaea pedrosoi in vitro .Conclusion:The screening data and bioactivity evaluation suggest the P.oryzae bioassay is suitable for preliminary screening of bioactive agents from marine organisms.

OBJECTIVETo investigate gene diagnosis of occult micrometastasis in the mediastinal lymph node in patients with non-small cell lung carcinoma (NSCLC) and to evaluate its prognostic significance.

METHODSWith mRNA expression of mucoid1 (MUC1) gene examined by RT-PCR, 168 mediastinal lymph nodes taken from 37 pN(0) (negative lymph nodes) NSCLC patients (stage Ia approximately IIb) made up the experiment group. Thrity negative lymph nodes from 14 benign lesions and 30 positive lymph nodes from 15 NSCLC patients served as control. The survival difference between MUC1 mRNA-negative and MUC1 mRNA-positive groups was compared by the chi(2) test.

RESULTSUC1 mRNA was not identified in the negative-control group (specificity = 100%), but it was identified in 26 of 30 positive-control samples (sensitivity = 86.7%). MUC1 mRNA was identified in 16 (9.5%) of the experiment group from 12 patients whose TNM stage was up-regulated to stage IIIa. The 3-year survival rate (58.3%) of MUC1 mRNA positive group patients with occult micrometastasis in mediastinal lymph node was lower than the 88.0% of MUC1 mRNA negative group (P < 0.05).

CONCLUSIONOccult micrometastasis in the mediastinal lymph node in NSCLC patients can be diagnosed by MUC1 mRNA expression through RT-PCR. Poor prognosis in some pN(0) NSCLC patients may be associated with nodal occult micrometastasis.

Adult ; Aged ; Carcinoma, Non-Small-Cell Lung ; diagnosis ; secondary ; Female ; Genetic Markers ; genetics ; Humans ; Lung Neoplasms ; diagnosis ; pathology ; Lymphatic Metastasis ; diagnosis ; Male ; Middle Aged ; Mucin-1 ; analysis ; genetics ; Prognosis ; RNA, Messenger ; analysis

Objective To evaluate the virulence of NTV modified strain NTV-C7L in vitro and in vivo.Methods The non-replicative vaccinia virus modified strain NTV-C7L was constructed by homologous recombination,in order to evaluate the virulence of NTV-C7L by cell diffusion ability and virus replication,intranasal challenge,and scratch of tail of mice.Results The homologous recombination non-replicating vaccinia virus modified strain NTV-C7L was selected by blue-white screening,the purity of NTV-C7L was identified by PCR and Western blot,the C7L gene was successfully inserted and expressed;In human embryonic lung fibroblasts (MRC-5) as well as mouse embryonic fibroblasts (BALB/C3T3),the replication ability of NTV-C7L was recovered compared with that of NTV,but was lower than that of VTT;the result of the intranasal route with 106 PFU vaccinia virus showed that the animals infected with TTV had slightly greater weight loss than NTV or NTV-C7L(t =6.56,P＜0.001;t =5.73,P＜0.001).Compared with the NTV-C7L group,the weight loss of the NTV group was not statistically significant (t =0.597,P =0.081);the intranasal route with 107pFU vaccinia virus showed that the weight loss of the NTV group and NTV-C7L group was statistically significant compared with the TTV group (t =12.86,P＜0.001;t =10.71,P＜ 0.001).Compared with the NTV-C7L group,the weight loss of the NTV group was statistically significant(t =4.616,P＜0.001);the result of skin scratch indicated that mice scarified with TTV (106 PFU) formed more obvious skin redness,swelling and injury on the post infected 5 days,whereas only redness was detected after scarification with a much higher dose (107) of NTV or NTV-C7L.Conclusions The intracellular and mouse virulence of NTV modified strain NTV-C7L was recovered compared with NTV,but was significantly lower than that of TTV,which provided reference for optimization and application of vaccinia virus vector.

Objective:Using clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR associated protein 9 (Cas9) technology to edit Vaccinia virus (VACV) thymidine kinase (TK) Region for targeted recombination to establish an efficient vaccinia virus insertion recombination technology.Methods:We designed and synthesized guide RNAs(gRNA)targeting the TK region and then cloned individual gRNA into the PX458 vector that removes nuclear localization signals, and modified the original TK region recombinant plasmid. The gRNA and Cas9 co-expression plasmids were transfected into 293T cells separately to mediate the homologous recombination of vaccinia virus (VACV) and TK region recombination plasmid, and then the rate of viral recombination was evaluated by the appearance of blue and white spots.Results:The recombination efficiency mediated by the gRNA sequence designed and synthesized in this study is more than 1%, which is more than 10 times higher than the classical homologous recombination method .Conclusions:This study used CRISPR/Cas9 technology to establish a highly efficient recombinant vaccinia virus system, which provides technical support for pre-clinical research in vaccines or multivalent research of emerging infectious diseases, as well as tumor treatment.

Objective:To accurately ascertain the whole genome sequencing and the composition of open reading frames (ORFs) of non-replicating Tiantan strain of vaccinia virus (NTV) using next-generation long-read sequencing technology.Methods:NTV, obtained from our laboratory stock, was amplified and purified on chicken embryo fibroblast cells(CEFs), and the full-length genomic nucleic acid of NTV was extracted. The PacBio HiFi sequencing platform was utilized for de novo assembly to obtain the complete genomic sequence of NTV. Using a homology annotation strategy, we identified its ORF composition and compared it with known non-replicating vaccinia virus strains. Results:The total length of NTV′s genome was 171 729 bp, with a GC content of 33%. Its unique inverted terminal repeat (ITR) region comprised hairpin structures, two tandem repeat regions, and three non-repeat regions. NTV contained 166 ORFs, with major differences observed in the ITR and its surrounding regions when compared to MVA-BN and NYVAC. These three strains shared a common set of 138 ORFs. NTV encoded six unique ORFs related to virus evasion of host antiviral response.Conclusions:This study accurately determines the whole genome sequencing and ORFs composition of NTV, and reveals its similarities and differences with other replication-deficient vaccinia virus strains, which pave a way for the development and application of the next generation of monkeypox vaccines and novel viral vectors.

Parkinson disease （PD） is the second most prevalent neurodegenerative disorder and predominantly impacts the extrapyramidal system. This condition arises from the degeneration of the dopaminergic nigrostriatal pathway. In recent years， studies have shown that PD pathology and lesions are not limited to the center nervous system， and that PD is a systemic and multisystem disease. The concurrence of PD with peripheral neuropathy （PN） has been increasingly acknowledged， although its pathogenesis remains elusive. The motor symptoms in PD patients often mask the symptoms of PN， leading to relatively low clinical recognition of PD coexisting with PN. This poses challenges in the clinical diagnosis and treatment of PD. This review comprehensively summarize the pathogenesis of PD coexisting with PN.
Parkinson Disease

Objective To discover the medicinal active molecules from the fermentation extract of sponge-symbiotic Streptomyces sp. LHW2432. Methods Compounds were isolated and purified from the fermentation extract of LHW2432 by silica gel, ODS chromatographic columns, and HPLC. The structures of the compounds were elucidated based on the analyses of modern spectrum technologies and the related literatures research. Through plate coating method and broth microdilution method, the antimicrobial activities were tested by the indicator strains of Bacillus mycoides, methicillin-resistant Staphylococcus aureus (MRSA), Mycobacterium smegmatis, Candida Albicans, and Escherichia coli. Results Five compounds were discovered and their structures were identified as descycloavandulyl-lavanduquinocin (1), N-acetyltyramine (2), phomapyrone C (3), germicidin A (4), and germicidin I (5). Compound 1 showed inhibitory activities against MRSA (MIC, 100 μg/ml) and M. smegmatis (MIC, 64 μg/ml), respectively. Conclusion Five compounds were discovered from LHW2432, among which compound 1 was a new natural product and could be used as a precursor of the tricyclic carbazole alkaloids with neuroprotective activity. Moreover, compound 1 showed weak inhibitory activities against gram-positive pathogenic bacteria.

Objective To study the cyclic peptides from sponge Reniochalina sp. under the guidance of mass spectrometry. Methods Mass spectrometry-guided procedural separation methods were used to track and isolate the cyclic peptides from the sponge genus Reniochalina. The structures of compounds were elucidated by the determination of physicochemical parameters and comparison of spectroscopic data. The preliminary cytotoxic activity of compounds was assessed by the Cell Counting Kit-8 (CCK-8) method. Results Three cyclic peptides were isolated from the sponge Reniochalina sp. and identified as stylopeptide 1 (1), hymenamide D (2) and axinastatin 2 (3). Compound 1 exhibited cytotoxicity against six human cancer cell lines with IC₅₀ values ranging from 6.09 to 17.26 μmol/L. Conclusion Compound 1 - 3 were isolated from Reniochalina sp. for the first time, and compound 1 was a cytotoxic cyclic heptapeptide.