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The glycan loop of Zika virus (ZIKV) envelope protein (E) contains the glycosylation site and has been well documented to be important for viral pathogenesis and transmission. In the present study, we report that deletions in the E glycan loop, which were recorded in African ZIKV strains previously, have re-emerged in their contemporary Asian lineages. Here, we generated recombinant ZIKV containing specific deletions in the E glycan loop by reverse genetics. Extensive in vitro and in vivo characterization of these deletion mutants demonstrated an attenuated phenotype in an adult A129 mouse model and reduced oral infections in mosquitoes. Surprisingly, these glycan loop deletion mutants exhibited an enhanced neurovirulence phenotype, and resulted in a more severe microcephalic brain in neonatal mouse models. Crystal structures of the ZIKV E protein and a deletion mutant at 2.5 and 2.6 Å, respectively, revealed that deletion of the glycan loop induces encephalitic flavivirus-like conformational alterations, including the appearance of perforations on the surface and a clear change in the topology of the loops. Overall, our results demonstrate that the E glycan loop deletions represent neonatal mouse neurovirulence markers of ZIKV. IMPORTANCE Zika virus (ZIKV) has been identified as a cause of microcephaly and acquired evolutionary mutations since its discovery. Previously deletions in the E glycan loop were recorded in African ZIKV strains, which have re-emerged in the contemporary Asian lineages recently. The glycan loop deletion mutants are not glycosylated, which are attenuated in adult A129 mouse model and reduced oral infections in mosquitoes. More importantly, the glycan loop deletion mutants induce an encephalitic flavivirus-like conformational alteration in the E homodimer, resulting in a significant enhancement of neonatal mouse neurovirulence. This study underscores the critical role of glycan loop deletion mutants in ZIKV pathogenesis, highlighting a need for global virological surveillance for such ZIKV variants.

This study analyzed the seasonal variation, sources, and source-specific health risks of PM

Thes protocols describe the experimental methods in “S100A4+ Macrophages Assist Zika Virus to Invade and Persist in Seminiferous Tubules via Interferon-gamma Mediation”.

Nitrogenous species, as important chemical components in PM2.5, include organic nitrogen (ON) and inorganic nitrogen (IN), both of which have potential effects on human health, climate change and visibility degradation. In this study, we analyzed total nitrogen (TN) by CHN Elemental analyzer and inorganic nitrogen by ion chromatography (IC) respectively to obtain ON by calculating the difference between TN and IN. The results show that the mean ON concentrations in winter and summer are both 2.86 μg·m−3, ten times higher than other places reported on average. ON contributes about 20%–30% to TN on average in both seasons, presenting higher contribution in summer. N:C ratios are much higher in summer than winter. ON sources or formation were strengthened by heavy PM2.5 pollution loads, especially sensitive to sulfate. ON concentrations are higher at night in the both seasons, however with distinguished day and night difference patterns influenced by relative humidity (RH) conditions. In winter, ON concentrations increase with RH on average through low RH values to high RH values. The variations are far larger than the ones caused by day and night difference. However in summer, day and night difference dominates the variations of ON concentrations at low RH values, and RH conditions promote ON concentrations increase significantly only at high RH values. Dust related source and anthropogenic emission related secondary source are identified as important sources for ON. At heavy pollution loads, ON sources are more of secondary formation, possibly strengthened by combination influence of RH and acidity increase.

Thieno[2,3-d]pyrimidinones were reported to act as potent anticancer agents; in this work, a series of new substituted thieno[2,3-d]pyrimidinone (6) were synthesized via the aza-Wittig reaction in satisfactory yields. The structures of these compounds were confirmed by elemental analysis, IR, 1H-NMR, and mass spectral data, and compound 6h was further analyzed by single crystal X-ray diffraction. Cytotoxic effect of all the compounds was carried out on human breast and lung cancer cell lines (MCF-7 and SPC-A-1, A459). Compound 6f exhibited the best inhibition activities against A459 with IC50 4.1 μM.

In 2014, Guangzhou City, South China, suffered from its worst outbreak of dengue fever in decades. Larval mosquito habitat surveillance was carried out by using android mobile devices in four study sites in May 2015. The habitats with larval mosquitoes were recorded as photo waypoints in OruxMaps or in videos. The total number of potential mosquito habitats was 342, of which 166 (49%) were found to have mosquito larvae or pupae. Small containers were the most abundant potential habitats, accounting for 26% of the total number. More mosquito larvae and pupae, were found in small containers than in other objects holding water, for example, potted or hydroponic plants (p < 0.05). Mosquito larvae were collected from all plastic road barriers, used tires, and underground water. Aedes albopictus larvae were found from small and large containers, stumps, among others. The overall route index (RI) was 11.3, which was 14.2 times higher than the grade C criteria of the National Patriotic Health Campaign Committee (NPHCC), China. The higher RIs were found from the bird and flower markets, schools, and underground parking lots. The results indicated that Android mobile devices are a convenient and useful tool for surveillance of mosquito habitats, and the enhancement of source reduction may benefit the prevention and control of dengue vector mosquitoes.

Some of the new compounds (IV) show good cytotoxicity against human breast and lung cancer cell lines, with compound (IVf) possessing the best inhibitory activity against lung cancer cell line (A459).

In the title compound C20H21Cl2N2O2S, the thienopyrimidine ring system is essentially planar. The crystal packing is stabilized by van der Waals forces and by an intermolecular C—H...O hydrogen-bonding interaction, which links the molecules into zigzag chains.

In the crystal structure of the title compound, C22H21N3O2, the packing of the molecules is mainly governed by intermolecular C—H...O hydrogen bonds and π–π interactions.

The title compound, C19H21N3O3S, was synthesized via the aza-Wittig reaction of functionalized iminophosphorane with phenyl isocyanate under mild conditions. In the molecule, the fused thienopyrimidine ring system is essentially planar, with a maximum deviation of 0.072 (2) Å, and makes a dihedral angle of 60.11 (9)° with the phenyl ring. An intramolecular C—H...O hydrogen bond is present. The crystal packing is stabilized by intermolecular N—H...O and C—H...O hydrogen bonds.

Summary The aza-Wittig reactions of iminophosphorane 3 with aromatic isocyanates generated carbodiimides 4, which were reacted with alkylamines under mild conditions to give a series of 2-(alkylamino)-5,6-dimethylthieno[2,3-d]pyrimidin-4(3H)-ones 6 and 8 in satisfactory yield. Their structures were confirmed by 1H NMR, EI-MS, IR and elementary analysis, and compound 8c was further analyzed by single-crystal X-ray diffraction. The preliminary bioassays indicated that these compounds showed excellent fungicidal activities against six kinds of fungi.

MASPIN gene is closely associated with carcinogengsis and plays a key role in cell proliferation, adhesion, migration and apoptosis. This study was to construct a recombinant eukaryotic vector expressing MASPIN, and explore the effect of MASPIN overexpression on the apoptosis in human gastric carcinoma cell line SGC7901.A eukaryotic expression vector MASIPIN/PCR2.1 was constructed and transfected into SGC7901 cells. RT-PCR and Western blot were used to detect the expression changes of MASPIN and Bax/Bcl-2. TNF-related apoptosis inducing ligand (TRAIL) (50ng/ml) was used to induce apoptosis at different time courses. DNA apoptotic ladders were determined using agarose gel electrophoresis (AGE). Cell apoptosis was measured by flow cytometry (FCM).Recombinant plasmid MASIPIN/PCR2.1 was successfully constructed and transfected into SGC7901 cells. The mRNA and protein levels of MASPIN were significantly higher in the MASPIN/PCR2.1 group (33.6+/-1.2, 23.4+/-1.6) than in the PCR2.1(15.0+/-1.5, 12.3+/-1.5)and the untreated group (13.7+/-2.0, 12.0+/-1.3) (P0.05). After transfection of MASIPIN/PCR2.1, DNA apoptotic ladders appeared in SGC7901 cells and the induction of apoptosis was in a time-dependent manner. The apoptosis rates were 8.0%, 16.3% and 25.8%in the MASPIN/PCR2.1 plus TRAIL group, 3.0%, 8.2%, 14.4% in the MASPIN/PCR2.1 group, and 4.1%, 9.8%,15.9% in the TRAIL group at 12, 24, and 48 h(P0.05). The expression levels of Bax mRNA and protein at 48 h after MASIPIN/PCR2.1 transfection were significantly higher in MASPIN/PCR2.1 plus TRAIL group(55.3+/-2.1, 75.4+/-1.3) than in the PCR2.1 group (34.3+/-1.2, 40.7+/-1.8) and the TRAIL group (43.2 +/-1.8,36.2+/-1.3)(P0.05). The expression of Bcl-2 mRNA in the MASPIN/PCR2.1 plus TRAIL group, PCR2.1 group and TRAIL group were 28.3+/-2.5, 34.3+/-1.2, 32.8+/-2.1, respectively (P0.05), and those of Bcl-2 protein were 17.4+/-1.5, 45.1+/-2.1, 42.8+/-1.5 in the three groups, respectively (P0.05).upregulation of MASPIN/PCR2.1 can significantly enhance the sensibility of gastric cancer cell line SGC7901 to the apoptosis inducer. This maybe related to the upregulation of Bax and downregulation of Bcl-2.

To study the genetic aberrations and their pathologic significance in follicular lymphoma (FL).Paraffin-embedded tissue samples of 55 cases of FL, 28 cases of other small B-cell lymphomas and 10 cases of reactive follicular hyperplasia were retrieved. Nested polymerase chain reaction (PCR) was used to detect clonal rearrangement of immunoglobulin heavy chain gene (IgH) in FL and other small B-cell lymphomas. The translocation t (14; 18) was studied by PCR and dual-color fluorescence in-situ hybridization (FISH) in FL. Cases of reactive follicular hyperplasia were used as controls.Amongst the 55 cases studied, 49 cases were nodal and 6 cases were extranodal. There were 33 males and 22 females. The male-to-female ratio was 1.5:1. The median age of the patients was 57 years. Twenty-five cases belonged to histologic grade 1, while 19 cases were grade 2 and 11 cases were grade 3. Beta-actin DNA was detected in 50 cases of FL. Amongst those 50 cases, clonal IgH rearrangement was present in 34 (68%). Twenty-four cases (48%) and 25 cases (50%) were positive for FR3A and FR2 respectively. Fifteen cases (30%) showed dual positivity for both FR3A and FR2. Thirty-four cases (68%) demonstrated clonal IgH rearrangement. As for other small B-cell lymphomas, 25 cases were positive for beta-actin. FR3A and FR2 were detected in 18 and 17 cases respectively. Clonal IgH rearrangement was demonstrated in 24 cases. In contrast, none of the 4 cases of reactive follicular hyperplasia showed the clonal rearrangement pattern. Amongst the 44 cases of nodal FL analyzed, t (14; 18) was detected in 15 cases (with 14 cases in MBR and 1 case in mcr). In general, FISH was superior to PCR in detecting t (14; 18) using paraffin-embedded tissue samples.The detection rate of clonal IgH rearrangement in FL is lower than that in other small B-cell lymphomas. Demonstration of t (14; 18) in paraffin-embedded tissue samples by FISH helps in diagnosis of FL. FISH is superior to PCR, as the technique is more sensitive and less labor intensive.

To investigate bcl-6 protein expression and gene rearrangement patterns in diffuse large B-cell lymphoma (DLBCL) and their clinicopathologic significance.Immunohistochemical studies for bcl-6 and CD10 proteins were performed on 51 cases of DLBCL paraffin-embedded tissues (including 22 nodal samples and 29 extranodal samples) and 10 cases of reactive lymphoid hyperplasia (RLH) paraffin-embedded tissues. Interphase fluorescence in-situ hybridization (FISH) with dual color breakapart probe was also used to identify rearrangement of bcl-6 gene in 32 cases of nodal DLBCL tissues (including 22 paraffin-embedded samples and 10 fresh samples) and 5 cases of RLH paraffin-embedded tissues.(1) The rates of bcl-6 protein expression in nodal DLBCL, extranodal DLBCL and RLH were 72.7% (16/22), 75.9% (22/29) and 100.0% (10/10) respectively. The rates of CD10 expression were 40.9% (9/22), 41.4% (12/29) and 100.0% (10/10) respectively. All lymphoma samples which expressed CD10 also showed co-expression of bcl-6 protein. (2) The co-expression of bcl-6 and CD10 was observed in 40.9% (9/22) nodal DLBCL and 41.4% (12/29) extranodal DLBCL. Low clinical stage (stage I and II) was more frequently observed in cases with co-expression of bcl-6 and CD10 (P0.05). (3) The rates of bcl-6 gene rearrangement in nodal DLBCL was 28.1% (9/32), with 27.3% (6/22) in paraffin-embedded tissues and 30.0% (3/10) in fresh tissues. There was no statistically significant difference found between the two groups (P0.05). Bcl-6 gene rearrangement was not found in all the 5 cases of RLH, and there was a significant difference between RLH and DLBCL (P0.05).The rate of bcl-6 protein expression is high in DLBCL cases, and the detection of bcl-6 and CD10 protein co-expression may help in the diagnosis and differential diagnosis of DLBCL. Those DLBCL cases with co-expression of bcl-6 and CD10 may also have a better prognostic implication. On the other hand, bcl-6 gene rearrangement can be identified by interphase FISH with dual color breakapart probe in both paraffin-embedded and fresh lymphoma tissues.

A simple, sensitive and convenient ion chromatography(IC) method was established for the simultaneous determination of twelve water-soluble inorganic anions(F- , Cl- , NO2(-), NO3(-), SO3(2-), SO4(2-) , PO4(3-)), and fifteen water-soluble organic ions(formate, acetate, MSA, oxalate, malonate, succinate, phthalates, etc.) in atmospheric aerosols. The linear concentrations ranged from 0.005 microg/m3 to 500 microg/m3 ( r = 0.999-0.9999). The relative standard deviation (RSD) were 0.43%-2.00% and the detection limits were from 2.7 ng/m3 to 88 ng/m3. The proposed method was successfully applied to the simultaneous determination of those inorganic ions and organic ions in PM2.5 of Beijing.

To investigate BCL-6 gene mutations in B-cell non-Hodgkin lymphomas (B-NHL) and their implications in lymphoma pathogenesis.Polymerase chain reaction (PCR) and direct DNA sequencing methods were used to identify mutations in the 5'-noncoding region of BCL-6 gene in 135 cases of B-NHL, 5 cases of T-NHL, 5 cases of nodular lymphocyte predominance Hodgkin's lymphoma (NLPHL) and 10 cases of reactive hyperplasia of lymph node.Mutations were identified in 6 cases of nodal DLBCL (27.3%), 4 cases of FL (22.2%), 4 cases of MALT lymphoma (22.2%), 4 cases of extranodal DLBCL (20.7%) and 2 cases of LRH (20%). No mutations were detected in T-NHL and NLPHL (P0.05). There were no significant differences in incidences of BCL-6 gene mutations between nodal and extranodal DLBCL (P0.05). All mutations were base substitutions and the frequency of single-base change was 0.14 x 10(-2)/bp approximately 0.68 x 10(-2)/bp.Mutations of the 5'non-coding region of BCL-6 gene may be involved in the pathogenesis and progression of B-NHL. Molecular demonstration of such mutations may provide a marker of lymphomas derived from the germinal center-related B cells.

AIM: To investigate the significance of c-kit gene mutation in gastrointestinal stromal tumors (GIST). METHODS: Fifty two cases of GIST and 28 cases of other tumors were examined. DNA samples were extracted from paraffin sections and fresh blocks. Exons 11, 9 and 13 of the c-kit gene were amplified by PCR and sequenced. RESULTS: Mutations of exon 11 were found in 14 of 25 malignant GISTs (56%), mutations of exon 11 of the c-kit gene were revealed in 2 of 19 borderline GISTs (10.5%), and no mutation was found in benign tumors. The mutation rate showed significant difference (χ2 = 14.39, P < 0.01) between malignant and benign GISTs. Most of mutations consisted of the in-frame deletion or replication from 3 to 48 bp in heterozygous and homozygous fashions, None of the mutations disrupted the downstream reading frame of the gene. Point mutations and frame deletions were most frequently observed at codons 550-560, but duplications were most concentrated at codons 570-585. No mutations of exons 9 and 13 were revealed in GISTs, Neither c-kit gene expression nor gene mutations were found in 3 leiomyomas, 8 leiomyosarcomas, 2 schwannomas, 2 malignant peripheral nerve sheath tumors, 2 intra-abdominal fibromatoses, 2 malignant fibrous histiocytomas and 9 adenocarcinomas. CONCLUSION: C-kit gene mutations occur preferentially in malignant GISTs and might be a clinically useful adjunct marker in the evaluation of GISTs and can help to differentiate GISTs from other mesenchymal tumors of gastrointestinal tract, such as smooth muscle tumors, schwannomas, etc.

To explore the clinicopathological, immunohistochemical and molecular genetic features of intra-abdomen extra-gastrointestinal stromal tumors (EGISTs) and their differential diagnosis.Nine cases of EGISTs from the abdominal cavity or retroperitoneum which were previously diagnosed as leiomyoma, leiomyoblastoma, or leiomyosarcoma etc. by a panel of antibodies such as CD117, CD34, alpha-SMA, MSA, desmin, S-100, and PGP9.5 from which five cases were detected for c-kit gene mutation.The tumors occurred in 5 men and 4 women, the age ranged from 38 to 72 years (mean 61.7 years). Four cases arose from the mesentery, two from omentum, two from retroperitoneum and one located at the hilus of the spleen. The size of tumors ranged from 5 cm to 23 cm (mean 12.9 cm) in diameter and the tumor cell components varied: mainly spindle cells (seven cases), epithelioid cells (one case), mixed cells (one case). Tumors expressed CD117 (8/9), CD34 (5/9), alpha-SMA (3/9), MSA (4/9), desmin (0), S-100 protein (1/9) and PGP9.5 (1/9). Of the five cases examined for heterozygous deletion mutation of 11 exon of the c-kit gene two were found positive. Two borderline cases showed long-term survival of 8 years and 11 years, respectively. In seven malignant cases, two showed adverse outcome, one survived 4 years without recurrence, two were lost in follow up and two new cases were still being in followed.GIST-type stromal tumors can also occur in the abdomen, most cases were borderline or malignant, tumor coagulative necrosis, mitosesor= 5 per 50 high-power fields and obvious nuclear atypia indicating malignancy. Differential diagnosis of EGIST including benign or malignant smooth muscle tumors, benign or malignant nerve sheath tumors etc.

In the title compound, C18H23N5O2, the triazolopyrimidine ring system is essentially planar, with a maximum displacement of 0.032 (2) Å, and forms a dihedral angle of 87.59 (15)° with the phenyl ring. In the crystal, molecules are linked by intermolecular C—H...O hydrogen bonds and C—H...π interactions into chains parallel to the c axis.

In the title compound, C21H21N3OS, the three fused rings of the 1-benzothieno[3,2-d]pyrimidine system are almost coplanar. The crystal packing is stabilized by π–π stacking inter­actions and inter­molecular C—H⋯O hydrogen bonds.

In the crystal structure of the title compound, C21H20N2O3, mol­ecules are stacked through inter­molecular π–π inter­actions. No significant hydrogen bonds are observed.

In the crystal structure of the title compound, C21H19N3OS, there are inter­molecular C—H⋯O hydrogen bonds. No π–π stacking inter­actions are observed.

In the title compound, C14H16ClN5O3, there is evidence for significant electron delocalization in the triazolyl system. Intramolecular C—H...O and intermolecular C—H...O and C—H...N hydrogen bonds stabilize the structure.

In the title compound, C17H23N3O2S, the central thieno­pyrimidine ring system is essentially planar. The cyclo­hexene ring, in which the four CH2 groups are disordered in a 3:1 ratio, adopts a half-chair conformation and the morpholine ring is in a standard chair conformation. The mol­ecular structure is stabilized by intra­molecular C—H⋯O and C—H⋯N hydrogen bonds. In the crystal packing, inter­molecular C—H⋯O hydrogen-bonding inter­actions link the mol­ecules into chains along the b axis.

A tandem aza-Wittig reaction of ethyl 3-triphenyl­phospho­ranylideneamino-4-cyano-1-phenyl-1H-pyrrole-2-carboxyl­ate with 4-chloro­phenyl isocyanate and diisobutyl­amine produced the title compound, C27H28ClN5O. In the mol­ecule, the angles between the fused-ring system and the two benzene rings are 51.66 (2) and 65.79 (2)°. Intra­molecular C—H⋯N hydrogen bonds determine the conformation of the mol­ecule. The packing of the mol­ecules in the crystal structure is governed mainly by inter­molecular C—H⋯N hydrogen-bonding inter­actions. There are no π–π inter­actions.