Progress of miRNA-27a in gastric cancer / 肿瘤研究与临床

miRNA-27a (miR-27a) is a member of miR-23a-27a-24 gene cluster and is abnormally expressed in gastric cancer. miR-27a plays an important role in gastric cancer cell epithelial-mesenchymal transition, proliferation, microenvironment and resistance to chemotherapy drugs by regulating SFRP1, PHLPP2, BTG2, SOCS6, FOXO1 and other tumor suppressor genes. At the same time, single nucleotide polymorphisms of miR-27a and gastric cancer have also become hotspots in recent years. A more comprehensive exploration of the relationship between miR-27a and gastric cancer is expected to provide new methods and ideas in the treatment of gastric cancer. This article reviews the research progress of miR-27a in gastric cancer.

Target-specific gene delivery in plant systems and their expression: Insights into recent developments

High-yielding Indian cotton varieties are not amenable for regeneration and transformation because they arerecalcitrant in nature. In this work, we have developed Narasimha (NA1325) cotton variety by introducing threeCrygenes driven by three different promoters conferring insect resistance. The meristematic region of embryo axisexplants were infected and co-cultivated with Agrobacterium tumefacience (LBA4404) harbouring pMDC100vector with three Cry gene cassettes (a-globulin : Cry2Ab, DECaMV35s : Cry1F and nodulin : Cry1Ac) with Npt IIas a selectable marker gene. Out of 1010 embryo axes explants infected, 121 (T0) regenerated under two rounds ofkanamycin selection medium. About 2551 T1 seeds were collected from 111 T0 plants and these seeds screened againwith kanamycin at seedling stage. The transgenic plants were characterized by PCR, real time quantitative PCR,lateral flow strip protein assay and insect bioassay. Out of 145 kanamycin resistant plants (T1), twelve showedamplification of all four transgenes: Npt II, Cry2Ab, Cry1F and Cry1Ac through PCR with expected amplicons as395, 870, 840 and 618 bp, respectively. Further, lateral flow strip test revealed Cry1F and Cry1Ac proteinsaccumulated in 12 plants, whereas Cry2Ab protein was detected in eight only. The transcripts of all three Cry geneswere accumulated significantly higher in transgenic plants at T2 generation. The transgenic lines showed effectiveresistance against Helicoverpa armigera and Spodoptera litura larvae. The T2 line L-3 exhibited highest percentageof insect mortality, in which transcripts of all cry genes were accumulated higher than other plants. The transgeniccotton plants carrying triple Cry genes could be an excellent germplasm resource for the breeders for introgressions.

The phenotype and genotype characteristics of 41 patients with steroid 5α-reductase type 2 deficiency / 中华内分泌代谢杂志

Objective To investigate the cliuical phenotype and the genotype of forty-one patients with steroid 5α-reductase type 2 deficiency.Methods The clinical data were collected including physical examination,medical history,laboratory test,as well as ultrasonic examination.Genomic DNA was extracted from peripheral blood leukocytes.Sanger sequencing and targeted gene captured next-generation sequencing were applied to detect the SRDSA2 gene mutation.Results All the patients are Han nationality and their ages ranged from 4 months to 11 years old.The karyotypes of 41 patients were 46,XY and all SRY genes were detected as positive.There were 26 (63％) patients manifested isolated micropenis,and the rest of fifteen patients were hypospadias associated with microphallus accounting for 37％.There were 39 patients who carried biallelic mutation.Two cases just identified one allele mutation.Sixteen gene mutation types were confirmed.Among them c.725A ＞ G (p.Tyr242Cys),c.694C ＞ G (p.His232Asp),and c.548-9T＞G are the novel gene types.The allele frequency of c.680G＞A (p.Arg227Gln) is 60％ (48/80).Conclusion The primary manifestations of patients with steroid 5α-reductase type 2 deficiency were micropenis or hypospadias accompanied with micropenis.c.680G＞A (p.Arg227Gln) is the predominantly mutation type of Chinese patient with steroid 5α-reductase type 2 deficiency.

A New Integrated Newborn Screening Workflow Can Provide a Shortcut to Differential Diagnosis and Confirmation of Inherited Metabolic Diseases

PURPOSE: We developed a new workflow design which included results from both biochemical and targeted gene sequencing analysis interpreted comprehensively. We then conducted a pilot study to evaluate the benefit of this new approach in newborn screening (NBS) and demonstrated the efficiency of this workflow in detecting causative genetic variants. MATERIALS AND METHODS: Ten patients in Group 1 were diagnosed clinically using biochemical assays only, and 10 newborns in Group 2 were diagnosed with suspected inherited metabolic disease (IMD) in NBS. We applied NewbornDiscovery (SD Genomics), an integrated workflow design that encompasses analyte-phenotype-gene, single nucleotide variant/small insertion and deletion/copy number variation analyses along with clinical interpretation of genetic variants related to each participant's condition. RESULTS: A molecular genetic diagnosis was established in 95% (19/20) of individuals. In Group 1, 13 and 7 of 20 alleles were classified as pathogenic and likely pathogenic, respectively. In Group 2, 11 and 6 of 17 alleles with identified causative variants were pathogenic and likely pathogenic, respectively. There were no variants of uncertain significance. For each individual, the NewbornDiscovery and biochemical analysis results reached 100% concordance, since the single newborn testing negative for causative genetic variant in Group 2 showed a benign clinical course. CONCLUSION: This integrated diagnostic workflow resulted in a high yield. This approach not only enabled early confirmation of specific IMD, but also detected conditions not included in the current NBS.

The analysis of clinical manifestations and genetic mutations in a child with hereditary fructose intolerance / 临床儿科杂志

Objective To analysis the clinical and gene mutation characteristics of hereditary fructose intolerance (HFI). Methods The clinical features and the results of gene testing in the child with HFI and her parents were analyzed retrospectively. Gene sequencing was carried out by high-throughput sequencing and validated by Sanger sequencing. Results The 4-year-3-month old girl had recurrent hypoglycemia episodes and growth retardation. When the condition was stable, the levels of lactic acid and urine micro protein were slightly higher, and the levels of thyroid hormone, cortisol, glycosylated hemoglobin, insulin and C peptide were normal.EEG showed epileptiform activity.Gene sequencing revealed the presence of aldolase B gene(ALDOB) compound heterozygous mutations, a novel splicing mutations (c.325-1G>A) in intron 3 and a frameshift mutation (c. 865delC;p.L289fs*10) in exon 8. Her father carries a frameshift mutation, and her mother carries a splicing mutation. Conclusion The diagnosis of HFI caused by ALDOB mutation can be confirmed by high-throughput sequencing technology.

Targeted nano-vector for gene delivery into human bone marrow mesenchymal stem cells and cellular MR imaging in vitro / 中国医学影像技术

Objective To explore the feasibility and efficacy of an MRI-visible,targeted,nano-vector which is synthesized by attaching a targeting ligand,the GD2 single chain antibody (scAb GD2),to the distal ends of PEG-g-PEI-SPION as a carrier for gene delivery into human bone marrow mesenchymal stem cells (hBMSCs) and in vitro cellular MR imaging.Methods scAbGD2-PEG-g-PEI-SPION was synthesized as previously reported.Gel electrophoresis was performed to assess the pDNA condensation ability of scAbGD2-PEG-g-PEI-SPION.The particle size and Zeta potential of scAbGD2-PEG-g-PEI-SPION/pDNA nanocomplexes were observed by dynamic light scattering.Cytotoxicity of scAbGD2-PEG-g-PEI-SPI-ON was evaluated by CCK-8 assay using hBMSCs.Gene transfection efficiency of scAbGD2-PEG-g-PEI-SPION in hBMSCs was quantified by flow cytometry,PEG-g-PEI-SPION,scAbGD2-PEG-g-PEI-SPION,scAbGD2-PEG-g-PEI-SPION+ free AbGD2 and scAbIgG2a-PEG-g-PEI-SPION group was established.The cellular internalization of scAbGD2-PEG-g-PEI-SPION/pDNA nanocomplexes was observed by confocal laser scanning microscopy and Prussian blue staining.MRI of scAbGD2-PEG-g-PEI-SPION was performed by cellular MRI scanning in vitro.Results scAbGD2-PEG-g-PEI-SPION condensed pDNA to form stable nanocomplexes of 80-100 nm in diameter and showed low cytotoxicity to hBMSCs.At the same N/P ratio,the transfection efficiency of scAbGD2-PEG-g-PEI-SPION group was significantly higher than those of other groups (P＜0.001).At the optimal N/P ratio of 20,scAbGD2-PEG-g-PEI-SPION/pDNA obtained the highest transfection efficiency of (59.60 ± 4.50) ％ in hBMSCs.Furthermore,hBMSCs labeled with scAbGD2-PEG-g-PEI-SPION showed sensitive low signal intensity on MRI T2/T2 *-weighted images in vitro.Conclusion scAbGD2-PEG-g-PEI-SPION is an efficient MRL visible targeted nano vector for gene delivery into hBMSCs.

Progress of microRNA-29b in tumors / 肿瘤研究与临床

MicroRNA (miRNA) is a kind of highly conserved microRNA molecules that regulate gene expression after transcriptional levels in eukaryotes. Numerous evidences indicate that miRNA can regulate malignant phenotypes of neoplasms by repressing the expressions of critical oncogenes or tumor suppressor genes, and abnormal expression of miRNA becomes one of the most important features of cancer. The expression of miR-29b was low in multiple types of tumors and its up-regulated expression will depress the expression of some target genes, which plays a crucial role in the occurrence and development of various neoplasms and is expected to serve as a biomarker for early diagnosis and prognosis assessment becoming a novel target of drug development and new treatment.

Targeted nano-vector for gene delivery into human bone marrow mesenchymal stem cells and cellular MR imaging in vitro / 中国医学影像技术

Objective To explore the feasibility and efficacy of an MRI-visible,targeted,nano-vector which is synthesized by attaching a targeting ligand,the GD2 single chain antibody (scAb GD2),to the distal ends of PEG-g-PEI-SPION as a carrier for gene delivery into human bone marrow mesenchymal stem cells (hBMSCs) and in vitro cellular MR imaging.Methods scAbGD2-PEG-g-PEI-SPION was synthesized as previously reported.Gel electrophoresis was performed to assess the pDNA condensation ability of scAbGD2-PEG-g-PEI-SPION.The particle size and Zeta potential of scAbGD2-PEG-g-PEI-SPION/pDNA nanocomplexes were observed by dynamic light scattering.Cytotoxicity of scAbGD2-PEG-g-PEI-SPI-ON was evaluated by CCK-8 assay using hBMSCs.Gene transfection efficiency of scAbGD2-PEG-g-PEI-SPION in hBMSCs was quantified by flow cytometry,PEG-g-PEI-SPION,scAbGD2-PEG-g-PEI-SPION,scAbGD2-PEG-g-PEI-SPION+ free AbGD2 and scAbIgG2a-PEG-g-PEI-SPION group was established.The cellular internalization of scAbGD2-PEG-g-PEI-SPION/pDNA nanocomplexes was observed by confocal laser scanning microscopy and Prussian blue staining.MRI of scAbGD2-PEG-g-PEI-SPION was performed by cellular MRI scanning in vitro.Results scAbGD2-PEG-g-PEI-SPION condensed pDNA to form stable nanocomplexes of 80-100 nm in diameter and showed low cytotoxicity to hBMSCs.At the same N/P ratio,the transfection efficiency of scAbGD2-PEG-g-PEI-SPION group was significantly higher than those of other groups (P＜0.001).At the optimal N/P ratio of 20,scAbGD2-PEG-g-PEI-SPION/pDNA obtained the highest transfection efficiency of (59.60 ± 4.50) ％ in hBMSCs.Furthermore,hBMSCs labeled with scAbGD2-PEG-g-PEI-SPION showed sensitive low signal intensity on MRI T2/T2 *-weighted images in vitro.Conclusion scAbGD2-PEG-g-PEI-SPION is an efficient MRL visible targeted nano vector for gene delivery into hBMSCs.

Progress of microRNA-29b in tumors / 肿瘤研究与临床

MicroRNA (miRNA) is a kind of highly conserved microRNA molecules that regulate gene expression after transcriptional levels in eukaryotes. Numerous evidences indicate that miRNA can regulate malignant phenotypes of neoplasms by repressing the expressions of critical oncogenes or tumor suppressor genes, and abnormal expression of miRNA becomes one of the most important features of cancer. The expression of miR-29b was low in multiple types of tumors and its up-regulated expression will depress the expression of some target genes, which plays a crucial role in the occurrence and development of various neoplasms and is expected to serve as a biomarker for early diagnosis and prognosis assessment becoming a novel target of drug development and new treatment.

Progression of secreted frizzled-related protein family in cancer / 国际肿瘤学杂志

Secreted frizzled-related protein (SFRP) can inhibit the expression of Wnt signaling pathway through Frizzled protein.The silencing of SFRP gene promoter methylation is associated with the occurrence and metastasis of many cancers such as colorectal cancer,gastric cancer,liver carcinoma,lung cancer and ovarian cancer.Several studies have found that SFRP gene has latent clinical value,which is expected to become the novel target for the gene diagnosis and treatment of cancer.

Folate-conjugated polyspermine for lung cancer-targeted gene therapy

Biodegradable polyamines have long been studied as potential recombinant viral gene vectors. Spermine (SPE) is an endogenous tetra-amine with excellent biocompatibility yet poor gene condensation capacity. We have previously synthesized a polyspermine based on SPE and poly(ethylene glycol) (PEG) diacrylate (SPE-alt-PEG) for enhanced transfection performance, but the synthesized SPE-alt-PEG still lacked specificity towards cancer cells. In this study, folic acid (FA) was incorporated into SPE-alt-PEG to fabricate a targeted gene delivery vector (FA-SPE-PEG) via an acylation reaction. FA-SPE-PEG exhibited mild cytotoxicity in both cancer cells and normal cells. FA-SPE-PEG possessed higher transfection efficiency than PEI 25 K and Lipofectamine(®) 2000 in two tested cancer cell lines at functional weight ratios, and its superiority over untargeted SPE-alt-PEG was prominent in cells with overexpressed folate receptors (FRs). Moreover, in vivo delivery of green fluorescent protein (GFP) with FA-SPE-PEG resulted in highest fluorescent signal intensity of all investigated groups. FA-SPE-PEG showed remarkably enhanced specificity towards cancer cells both in vivo and in vitro due to the interaction between FA and FRs. Taken together, FA-SPE-PEG was demonstrated to be a prospective targeted gene delivery vector with high transfection capacity and excellent biocompatibility.

Research progress on drugs targeting p53 / 中国药理学通报

With the understanding of the development and pro-gress of the cancer,the research of targeted cancer drug devel-opment reaches into a new era.p53 is an important tumor sup-pressor gene,the protein coded by p53 plays a critical role in tumor suppression mainly by inducing cell cycle regulation, DNA repair and apoptosis.Nowadays,p53 becomes a relatively attractive target for anti-cancer drug development and there are some drugs targeting p53,moreover,APR-246 which targets mutant p53 is in Phase II clinical trial.In addition,it facilitates drugs discovery programmes in the challenging area of protein-protein interactions and mutant protein conformational change. The review discusses the research progress of drugs which target p53 and elucidates the characteristics and mechanisms of these compounds.

Advances of miR-221/222 in tumors / 国际肿瘤学杂志

Because of the characteristics of microRNAs (miRNAs) and their important regulative roles in human tumor diseases,they may be novel approaches for gene target therapy.Most studies focus on finding critical regulator miRNAs of oncogenes or tumor suppressor genes.Recent researches show that the expression of miR-221 and miR-222 are abnormal,and they play different roles in different types of human tumors.The regulation of miR-221 and miR-222 may be a novel effective therapy for tumor.

Construction of targeted ultrasound contrast agent with anti-IgG monoclone antibody and the evaluation on the effectiveness of its targeted adhesion in vitro / 中华超声影像学杂志

Objective To construct targeted ultrasound contrast agent carried goat anti-mouse IgG antibody (UCA-IgG) and evaluate the effectiveness of its targeted adhesion using parallel plate flow chamber. Methods The ultrasound contrast agent targeted to mouse IgG was designed by conjugating monoclonal antibodies against mouse lgG to the lipid monolayer shell of the agent using biotin-streptavidin. The binding of IgG antibodies to the ultrasound contrast agent were identified by fluorescence in vitro. The attachment and detachment of UCA-IgG to mouse IgG immobilized on a culture dish were assessed in a parallel-plate flow chamber. While the plate lacked mouse IgG,or blocked with large number of goat anti-mouse IgG were served as two control groups. Results UCA-IgG issued a bright green fluorescence, while the contral lipid ultrasound contrast agent didn't show fluorescence. The number of UCA-IgG bound to mouse IgG of experimental group was greater than two control groups,increased with increasing coverslips surface antibody concentrations (P<0. 05),and there was significant positive correlation between the number of UCA-IgG bound to mouse IgG and time of combination (P<0.05). The adhesion rate of experimental group increased with shear stress before 0. 5×10-5 N/cm2 (P<0.05) and then decreased (P<0. 05). There was limited adherence of control groups to the UCA-IgG. The stess of half-maximal detachment was increased with increasing coverslips surface antibody concentrations (P<0.05). Conclusions UCA-IgG could adhere to mouse IgG in the physical conditions. It may provide strong supports for studying other targeted ultrasound contrast agent preliminary and fatherly in vitro.

Construction of pET15b-Z-VP1 vector for targeting gene delivery / 西安交通大学学报(医学版)

Objective To construct the vector pET15b-Z-VP1 by inserting the Z fragment into amino-terminal of JCV VP1.Methods The VP1 and Z fragment were amplified by PCR from plasmid pET15b and pEZZ18 respectively,and then they were linked by recombinant PCR.The Z-VP1 fragment was inserted into plasmid pET15b by restriction enzyme BamHⅠ and NcoⅠ.Results The VP1 and Z fragment were obtained by PCR and gel purification.The Z-VP1 fragment,which was linked by recombinant PCR from VP1 and Z fragment,was inserted into plasmid pET15b between BamHⅠ and NcoⅠ sites,and confirmed by enzyme digestion and DNA sequencing.The expression of VP1-Z was confirmed by Western blotting.Conclusion The plasmid pET15b-Z-VP1 has been constructed successfully by inserting Z fragment into amino-terminal of VP1.