Detection of pathogenic variants in four patients with globozoospermia / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic basis for 4 patients with globozoospermia.@*METHODS@#Semen and blood samples were collected from the patients for the determination of sperm concentration, viability, survival rate, morphology and acrosome antigen CD46. Meanwhile, DNA was extracted for whole exome sequencing (WES), and candidate variants were validated by Sanger sequencing.@*RESULTS@#All of the four patients were found to harbor variants of the DPY19L2 gene. Patients 1 ~ 3 had homozygous deletions of the DPY19L2 gene. Sanger sequencing confirmed that the DPY19L2 gene in patient 3 was disrupted at a recombination breakpoint area BP2, resulting in nonallelic homologous recombination and complete deletion of the DPY19L2 gene. Patients 2 and 3 respectively harbored novel homozygous deletions of exons 2 ~ 22 and exons 14 ~ 15. Patient 4 harbored heterozygous deletion of the DPY19L2 gene, in addition with a rare homozygous deletion of the 3' UTR region.@*CONCLUSION@#DPY19L2 gene variants probably underlay the globozoospermia in the four patients, which has fit an autosomal recessive pattern of inheritance and the characteristics of genomic diseases.

Expression and diagnostic value of miRNA-29c in type 1 diabetic patients with early nephropathy / 中华内分泌外科杂志

Objective:To study the expression of miRNA-29c in type 1 diabetic patients with early nephropathy and its diagnostic value for early nephropathy.Methods:168 patients with type 1 diabetes who were treated in our hospital from Jan. 2019 to Mar. 2022 were retrospectively selected as the research subjects. According to the occurrence of nephropathy, they were divided into simple diabetes group (122 cases) and diabetic nephropathy group (46 cases). Serum miRNA-29c levels were detected by RT-PCR. The gender, age, course of disease, and serum miRNA-29c levels were compared between the two groups. Logistic regression was used to analyze the influencing factors of early nephropathy in patients with type 1 diabetes. The ROC curve was used to analyze the diagnostic value of miRNA-29c for early nephropathy in type 1 diabetes.Results:The course of disease, blood pressure (systolic blood pressure, diastolic blood pressure), HbA1c, TC and blood uric acid in early nephropathy group were higher than those in simple diabetes group, while albumin, total bilirubin and miRNA-29c were lower than those in simple diabetes group, and the difference was statistically significant ( P<0.05). Multivariate Logistic analysis showed: long disease duration ( OR=2.061, 95% CI=1.090-3.896), systolic blood pressure ( OR=1.143, 95% CI=1.023-1.279), diastolic blood pressure ( OR=1.151, 95% CI=1.022) -1.298), high HbA1c ( OR=1.317, 95% CI=1.049~1.653), high blood uric acid ( OR=1.306, 95% CI=1.028-1.659), low miRNA-29c ( OR=0.845,95% CI= 0.730-0.979) were the risk factors for early nephropathy in patients with type 1 diabetes ( P<0.05). ROC curve analysis showed that the cut-off value of miRNA-29c for the diagnosis of early renal disease was 0.952, the area under the curve (AUC) was 0.863 (95% CI: 0.801-0.925), and the sensitivity and specificity were 84.78% and 80.33%, respectively. Conclusion:Serum miRNA-29c in patients with early stage nephropathy of type 1 diabetes is in a low expression state, which is an influencing factor for early stage nephropathy in patients with type 1 diabetes, and has a good diagnostic value for early stage nephropathy.

Genetic analysis of a pedigree affected with congenital high myopia caused by a novel splice site variant of COL11A1 gene / 中华医学遗传学杂志

OBJECTIVE@#To analyze genetic variant in a pedigree affected with congenital high myopia.@*METHODS@#Whole exome sequencing (WES) was carried out for the proband. Suspected variation was verified with Sanger sequencing. The pedigree was also subjected to co-segregation analysis.@*RESULTS@#WES has identified a novel splice site heterozygous variant (c.2556+1G>A) in the COL11A1 gene in the proband. Co-segregation analysis of the pedigree showed that the affected mother and two daughters of the proband have carried the same variant(c.2556+1G>A), while his unaffected father and sister did not. Based on the ACMG Standards and Guidelines for the Interpretation of Sequence Variants, the variant was classified as "likely pathogenic" (PVS1+PM2).@*CONCLUSION@#A novel splice variant (c.2556+1G>A) of the COL11A1 gene has been identified in a pedigree affected with congenital high myopia, which probably underlies the disease.

Genetic analysis of a pedigree affected with congenital high myopia caused by a novel splice site variant of COL11A1 gene / 中华医学遗传学杂志

Objective@#To analyze genetic variant in a pedigree affected with congenital high myopia.@*Methods@#Whole exome sequencing (WES) was carried out for the proband. Suspected variation was verified with Sanger sequencing. The pedigree was also subjected to co-segregation analysis.@*Results@#WES has identified a novel splice site heterozygous variant (c.2556+ 1G>A) in the COL11A1 gene in the proband. Co-segregation analysis of the pedigree showed that the affected mother and two daughters of the proband have carried the same variant(c.2556+ 1G>A), while his unaffected father and sister did not. Based on the ACMG Standards and Guidelines for the Interpretation of Sequence Variants, the variant was classified as "likely pathogenic" (PVS1+ PM2).@*Conclusion@#A novel splice variant (c.2556+ 1G>A) of the COL11A1 gene has been identified in a pedigree affected with congenital high myopia, which probably underlies the disease.

Phenotypic and genetic analysis of an inv dup(15) case with a BP3:BP3 rearrangement / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To analyze a case of supernumerary marker chromosome (SMC) with combined genetic techniques and explore its correlation with the clinical phenotype.</p><p><b>METHODS</b>The SMC was analyzed with G-banded karyotyping, multiplex ligation dependent probe amplification (MLPA), fluorescence in situ hybridization (FISH), and single nucleotide polymorphism array (SNP-array).</p><p><b>RESULTS</b>G-banding analysis indicated that the patient has a karyotype of 47,XX,+mar. MLPA showed that there were duplications of proximal 15q. FISH assay using D15Z4 probes indicated that the SMC was a pseudodicentric chromosome derived from chromosome 15. And SNP-array revealed that there were two extra copies of 15q11-13 region spanning from locus 20 161 372 to 29 071 810.</p><p><b>CONCLUSION</b>The duplication of Prader-Willi/Angelman syndrome critical region probably underlies the abnormal phenotype of the inv dup(15) case with a BP3:BP3 rearrangement.</p>

Genetic diagnosis and analysis of related genes for a pedigree with 2p25 and 12p13 cryptic rearrangements / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To analyze chromosome aberration in a child with mental retardation and abnormalities and its parents.</p><p><b>METHODS</b>Chromosome G banding, multiplex ligation-dependent probe amplification, fluorescence in situ hybridization and single nucleotide polymorphisms array were employed for analysis.</p><p><b>RESULTS</b>Karyotype analysis revealed that the child was 46,XX and the father was 46,XY, while the mother was 46,XX, add (12)(p13). Subtelomeric region analysis with MLPA displayed that the child has reduced ACP1 gene copy number in 2p25 region and increased SLC6A12,KDM5A gene copy numbers in 12p11 region. SNP-array has fine mapped the duplication to 12p13.33-p12.3, a 15.142 Mb region, and a deletion to 2p25.3 for 3.194 Mb, which resulted in duplication of 9 genes including SLC6A12 as well as deletion of 11 genes including SNTG2, respectively. FISH analysis revealed that the child was 46,XX,ish,der(2),t(2;12)(p25;p13)mat, or partial monosomy 2p25 and partial trisomy 12p13. In addition,the mother was a carrier with cryptic balanced translocation chromosome, 46,XX,isht(2;12) (p25;p13). Mental abnormalities and retardation of the child may be attributed to heterozygous deletion of SNTG2, MYT1L genes and duplication of SLC6A12 gene.</p><p><b>CONCLUSION</b>Combined use of MLPA, FISH and SNP-array can facilitate accurate diagnosis of cryptic rearrangement at genomic level.</p>

A novel mutation of NTRK1 gene in a family with congenital insensitivity to pain with anhidrosis / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To screen for mutations in the neurotrophic tyrosine kinase receptor type 1 (NTRK1) gene in a Chinese family affected with congenital insensitivity to pain with anhidrosis (CIPA).</p><p><b>METHODS</b>With informed consent obtained, peripheral blood samples were obtained from the patient and his family members. Seventeen coding exons and intron-exon boundaries of the NTRK1 gene were amplified with PCR and analyzed by direct sequencing.</p><p><b>RESULTS</b>A novel mutation c.2086_2087insC (p.Arg696 fsx) was identified in exon 16 of the NTRK1 gene in the proband. This insertion has caused open reading frame shifting and a premature termination has occurred just one codon downstream. Truncation of 72 amino acids at the C terminus has wiped out part of the tyrosine kinase domain (TKD) of the protein. Both of the proband's parents and two grandmothers have carried the c.2086_2087insC (p.Arg696 fsx) mutation. No mutation was found in the NTRK1 gene of other siblings.</p><p><b>CONCLUSION</b>Mutation analysis of the NTRK1 gene has been carried out in a Chinese family affected with CIPA, and a novel NTRK1 gene mutation was identified.</p>

Analysis of APC gene mutation in a familial adenomatous polyposis pedigree / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To analyze mutation of adenomatous polyposis coli (APC) gene in a family affected with familial adenomatous polyposis.</p><p><b>METHODS</b>The diagnosis was made based on clinical manifestations, family history, presence of numerous polyps in the colon as well as pathological examination. Peripheral blood samples were collected, and genomic DNA was extracted. Potential mutation of the APC gene was detected by polymerase chain reaction (PCR) and DNA sequencing. After finding the mutation in the proband, the same mutation was screened among other family members. The mutation was also confirmed with PCR-restriction fragment length polymorphism (RFLP), with which 100 unrelated healthy controls were examined.</p><p><b>RESULTS</b>A novel heterozygous nonsense mutation c.2891T>G (L964X) of the APC gene was identified in this pedigree. The mutation has led to premature termination of translation. The same mutation was not detected among the 100 healthy controls.</p><p><b>CONCLUSION</b>The c.2891T>G (L964X) of the APC gene probably underlies the familial adenomatous polyposis in this pedigree. The combined DNA sequencing and PCR-RFLP method is efficient and accurate for the diagnosis.</p>

Analysis of small supernumerary marker chromosome 15q11 in four infertile males / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To delineate the origins of small supernumerary marker chromosomes (sSMCs) identified in 4 infertile males.</p><p><b>METHODS</b>The sSMCs were analyzed with combined G-banding, N-banding, multiplex ligation-dependent probe amplification (MLPA), fluorescence in situ hybridization (FISH) and single nucleotide polymorphisms array (SNP-array) techniques.</p><p><b>RESULTS</b>G-banding analysis has suggested a 46,X,-Y,+mar karyotype in all of the 4 cases. N-banding revealed that all of the sSMCs have possessed two satellites located on both sides. By MLPA, 1 patient showed copy number gains for 15q11.2 region. SNP-array analysis suggested that all had duplication for 15q11.1-q11.2 region, spanning 3.06 Mb, 0.9118 Mb, 1.728 Mb and 0.287 Mb, respectively. By FISH analysis, all of the sSMCs showed two hybridization signals, indicating that they were dicentric chromosomes.</p><p><b>CONCLUSION</b>In all of the four cases, the marker chromosomes have derived from chromosome 15 and were bisatellited and dicentric, which gave rise to a karyotype of 47,XY,+ish,inv dup(15)(q11)(D15Z4++). sSMC 15q11 therefore may be a major cause for male infertility.</p>

Subtle mutation analysis of survival motor neuron gene in families with spinal muscular atrophy / 中华检验医学杂志

ObjectiveTo establish a analytical system for the survival motor neuron (SMN) subtle mutation,and evaluate its application in two families with spinal muscular atrophy (SMA).MethodsSMN genes in seven family members from two SMA families were analyzed at both transcript level and genomic level,by the use of the conventional PCR-RFLP,allele-specific PCR,multiplex ligation-dependent probe amplification (MLPA) and T subcloning and sequencing of SMNI gene.ResultsIn family A,the patient had a single SMN1 copy who was carrying nonsense mutation L228X,which was also found in his father.In family B,as the patient's sample was unavailable,the father was indeed a carrier with one normal SMN1 allele and the other SMN1 allele carrying a frameshift mutation 22_23insA.The remaining family members were SMA carriers with one SMN1 copy.ConclusionThis analytical system for SMN subtle mutation offers viable molecular basis for genetic counseling and prenatal diagnosis in SMA families.

Prenatal molecular diagnosis of four fetuses at high risk for X-linked adrenoleukodystrophy / 中华妇产科杂志

Objective To investigate methods for prenatal molecular diagnosis of fetuses at high risk for X-linked adrenoleukodystrophy(X-ALD).Methods The amniotic fluid was obtained and genomic DNA was isolated from amniotic fluid cells.Maternal contamination was evaluated by paternity test.PCRRFLP,sequencing and denaturing high performance liquid chromatography(DHPLC)were used to detect the ABCD1 gene of fetal genome.Results In the pedigree 1,the PCR product(799 bp)of the fetus 1 and her father(normal control)could be digested with BcnI. No P560L mutation,which was present in the index patient,was detected in the ABCD1 gene from the genomic DNA of the fetus 1 using direct sequencing.In the pedigree 2,the PCR product(232 bp)of the fetus 2 and her father could not be digested with MaeI and no Q177X mutation,which was present in the propositus,was detected in the ABCD1 gene from the genomic DNA of the fetus 2 using direct sequencing.In the pedigree 3,the PCR product(271 bp)was digested with AciI.the pattern of the fetus 3 and the propositus being the same,and the R617C mutation was found in the ABCD1 gene from the genomic DNA of the fetus 3 using direct sequencing.In the pedigree 4,the PCR product(269 bp)was analyzed with the DHPLC,and the pattern of elution peaks of the fetus 4 and her father was similar,but different from that of the propositus.No K276E mutation was detectable in the ABCD1 gene from the genomic DNA of the fetus 4 by using direct sequencing.Judging from the sex of the fetuses,fetuses 1 and 2 were normal homozygotes,fetus 3 was an ALD hemizygote,and fetus 4 was a normal hemizygote.Conclusion A new protocol for X-ALD prenatal molecular diagnosis is proposed,which would ensure the accuracy of prenatal diagnosis.

Establishment of a cellular model with human NADH-cytochrome b5 reductase deficiency via RNA interference / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To establish a cell line with human NADH-cytochrome b5 reductase (b5R) deficiency via RNA interference (RNAi).</p><p><b>METHODS</b>Two siRNA expressing vectors targeting the b5R mRNA were designed and constructed. Hepatocellular carcinoma BEL-7402 cells were transiently transfected with the two recombinants by lipofectamine (TM) 2000, and semi-quantitative RT-PCR was carried out to analyze the suppression of b5R mRNA; BEL-7402 cells stably transfected with the two siRNA expressing vectors were selected in the media with G418. By analyses of the mRNA, enzymatic activity and protein level of b5R, several cell clones with deficiency of b5R were established. The cell growth curve of BEL-7402 cells with b5R deficiency was detected by MTT assay.</p><p><b>RESULTS</b>Two siRNA expressing vectors targeting b5R mRNA were obtained, namely pSib5R-1 and pSib5R-2. When BEL-7402 cells were transfected transiently with pSib5R-2, the expression of b5R mRNA was significantly suppressed with a suppression ratio of 68.3%, indicating that pSib5R-2 could trigger the degradation of b5R mRNA effectively. Eighteen clones stably integrated exogenous plasmids were obtained. In two clones from pSib5R-2 transfection, the expression of b5R mRNA was suppressed by up to 48.2% and 56.2%, and the enzymatic activity was inhibited by up to 54.6% and 63.5%, respectively. The protein levels also decreased significantly. The defect of b5R did not change the cell growth rate.</p><p><b>CONCLUSION</b>The expression of b5R in BEL-7402 could be suppressed by vector-based RNA interference effectively. We established a cellular model with defect of b5R successfully, which can be used as a tool in investigation of the biological function of b5R and molecular mechanism of type II recessive congenital methemoglobinemia.</p>

Expression and preliminary use of human sperm-specific lactate dehydrogenase in Escherichia coli / 中华检验医学杂志

Objective To construct a recombinant vector of sperm-specific human lactate dehydrogenase ( hLDH-C4 ), express it in Escherichia coli ( E. coli ) BL21 ( DE3 ) and utilize it in the detection of anti-sperm antibody. Methods The coding sequence of hLDH-C4 was amplified from human testis λTripIEx cDNA library, and inserted into pET-28a( + ) after restriction enzyme digestion with Hind Ⅲ and Xho Ⅰ. The resultant recombinant vector was used to transform E. coli BL21 ( DE3 ) and the His-Tag fused hLDH-C4 was expressed after induction with IPTG. Western blot was used to analyzed the recombinant protein and LDH activity of bacterial lysates was determined. An indirect ELISA method for the detection of anti-sperm antibody was established by using the recombinant hLDH-C4 as antigen matrix. Results pET-28a( + )-hLDHC was successfully established. The protein with size of 35kD could be induced by IPTG when the recombinant plasmid was transfected into E. coli BL21 ( DE3 ). Western blot showed that the recombinant protein could be specifically recognized beth by anti-His tag monoclonal antibody and by rabbit anti-human LDH-C4 antibody. In addition, the recombinant protein showed high-level LDH activity when the bacterial lysate after IPIG induction was used to check LDH activity. The recombinant hLDH-C4 was confirmed when it was used in indirect EL1SA to detect anti-hLDH-C4 antibody. Conclusions The coding sequence of hLDH-C4 is cloned into the vector pET-28a( + ) and recombinant hLDH-C4 was expressed at a high level in E. coli. The recombinant hLDH-C4 is useful in the detection of anti-sperm antibody.

Expression and characterization of subunit C of mouse lactate dehydrogenase in Escherichia coli / 中华男科学杂志

<p><b>OBJECTIVES</b>To construct a prokaryotic recombinant vector for mouse lactate dehydrogenase-C and to detect its expression in BL21.</p><p><b>METHODS</b>The coding sequence of mouse lactate dehydrogenase subunit C was amplified from mouse testis RNA with specific primers, and cloned into pGEX-2T after the restriction digestion with BamH I and EcoR I. GST fusion protein was expressed after induction with IPTG.</p><p><b>RESULTS</b>Sequencing and restriction digestion of the recombinant plasmid revealed the existence of coding sequence for mouse lactate dehydrogenase subunit C. A protein band of about 60,000 could be induced by IPTG in the recombinant plasmid.</p><p><b>CONCLUSIONS</b>The coding sequence of mouse lactate dehydrogenase subunit C was introduced into the pGEX-2T plasmid and a GST-fused protein could be induced at a high level.</p>

Method for molecular diagnosis of hereditary methemoglobinemia / 中华检验医学杂志

Objective To investigate molecular diagnostic method for hereditarymethemoblobinemia. Methods The cDNA coding sequence of NADH-cytochrome b5 reductase (b5R) from 3 patients with hereditary methemoglobinemia was analyzed by direct sequencing of RT-PCR products and the genomic DNA of b5R gene by PCR-restriction endonuclease digestion or PCR-sequencing. Results The b5R cDNA of patient A was T/C heterozygous at nucleotide 527 and G/A heterozygous at nucleotide 608. The b5R cDNA of patient B was G/A heterozygous at both nucleotide 170 and nucleotide 179. The b5R cDNA of patient C was G/A heterozygous at nucleotide 608 and C/T heterozygous at nucleotide 791. Result of genomic DNA analysis was in agreement with that of cDNA approach. Conclusion The method for molecular diagnosis of hereditary methemoglobinemia was established and 3 novel b5R gene mutations were identified in compound heterozygosity in 3 Chinese patients.

cDNA cloning and prokaryotic expression of human pancreatic alpha-amylase / 中华检验医学杂志

Objective To obtain recombinant human pancreatic alpha-amylase protein (AMY2A). Methods Human pancreatic AMY2A cDNA was synthesized by RT-PCR using total RNA from human pancreatic tissues and a couple of primers designed according to the know sequence of human pancreatic alpha-amy lase gene, then digested with BamH Ⅰ and Kpn Ⅰ and inserted into the prokaryotic expression plasmid pGEX-5T vector. Construct The prokaryotic expression vector pGEX-5T-AMY2A was constructed and transformed into E. coli BL21 cell . Protein expressed under the induction of IPTG. The inclusion bodies were isolated and solubilized with urea and washed denatured and refolded. The fusion protein wes purified by affinity chrom atography with glutathione agarose. Results Sequence and restrction analysis revealed AMY2A gene was cloned in frame into pGEX-5T, SDS-PAGE profile showed a clear protein band with a relative molecular weight of 84000 and western blot indicated that the expressed product specifically reacted to polyclonal anti -human pancreatic AMY2A genes. Conclusion Human pancreatic AMY2A gene was successfully cloned,expressed and purification.

Effect of IL-12 on IFN-gamma and IL-10 produced by peripheral blood mononuclear cells in patients with chronic hepatitis B virus infection during IFN-alpha treatment / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the effect of IL-12 on IFN-gamma and IL-10 production of peripheral blood mononuclear cells (PBMC) in chronic hepatitis B virus infection patients during IFN-alpha treatment.</p><p><b>METHODS</b>Before and after IFN-alpha treatment of 3 months and 6 months, PBMC of 20 patients with chronic hepatitis B virus infection were collected and cultured in vitro in the culture fluid containing PHA (100 microg/ml), HBcAg (1 microg/ml), or HBeAg (1 microg/ml) for 48 h under the condition of the presence or absence of IL-12 (10 ng/ml). Then the levels of IFN-gamma and IL-10 were determined by ELISA.</p><p><b>RESULTS</b>There were 12 responders and 8 nonresponders to IFN-alpha treatment. In the responders, the enhancing effect of IL-12 on IFN-gamma production was significantly greater after IFN-alpha treatment than before the treatment. The production of IL-10 was suppressed in the presence of IL-12 after 3 months and 6 months of IFN-alpha treatment. In the same treatment time, the level of IFN-gamma in the presence of IL-12 was significantly higher than that in the absence of IL-12. To the nonresponders, the enhancing effect of IL-12 on IFN-gamma production was also significantly increased after IFN-alpha treatment. Moreover, in the same treatment time, the level of IFN-gamma in the presence of IL-12 was significantly higher than that in the absence of IL-12.</p><p><b>CONCLUSIONS</b>The enhancing effect of IL-12 on IFN-gamma production of PBMC in patients with chronic hepatitis B virus infection is increased during IFN-alpha treatment. IFN-alpha and IL-12 may enhance the efficacy for the treatment of chronic hepatitis B virus infection.</p>

Gene-based individual identification of hardly recognizable victims in an accident / 解放军医学杂志

Objective To identify hardly recognizable victims of an accident. Methods Tissues of muscle and cartilage were obtained from the dead bodies. Some of the tissues were checked by routine pathological microscopy. Genomic DNA was isolated from the tissues and subjected to STR profiling of 16 sites via multiple fluorescent PCR analysis with ABI’s AmpFLSTR Amplification Kit. Individual identification of the victims was carries out by matching the STR profiles of the victims with those of the parents. Results Routine pathological microscopy showed that the structure of some of the muscle tissues was totally destroyed, while the structure of all cartilage tissues was basically intact. Three patterns of genomic DNA isolated from victims’ muscle tissues could be seen in gel electrophoresis, i.e. basically undegraded, partially degraded and totally degraded. STR profiling failed due to the degradation of genomic DNA of some of the muscle tissues, while all samples of the cartilage genomic DNA could be used for STR typing. Conclusion Paternity identification based on STR genotyping was an effective way to identify victims of accidents, and cartilage tissue from the victims was the first choice for that purpose.

Effect of insulin-like growth factor 1 receptor ?-strain interrupted on lung adenoma cell line SPC-A-1 / 中国癌症杂志

Purpose:To study the effect of insulin-like growth factor 1 receptor ?-strain(IGF1R-?) interrupted by a special antibody (IGFⅠR-?Mab)on lung adenoma cell line SPC-A-1.Methods:IGFⅠR-?Mab was extracted by hybrid technology. SPC-A-1 cells were separated into 2 groups,the IGFⅠR-?Mab and the blank control. The IGFⅠR-?Mab cells were interfered by different densities of IGFⅠR-?Mab, including 20,40,60,80,100,120,140,160,180 and 200 ng/ml. The MTT curve line, morphology, ultrastructure and cell cycles were observed at 0,24,48,72 hours after the intervention respectively. Results:Compared with the control, apoptosis in IGFⅠR-?Mab group was significant(P= 0.009)and proliferation rate was decreased obviously within 160 ng/ml. However, the proliferation rate was no significant when the special antibody density was more than 200 ng/ml.Conclusions:The affinity of IGFⅠR-?Mab at IGF1R ?-strain is high. The interruption of IGF1R ?-strain by IGFⅠR-?Mab shows the obvious biological effects in vitro ,with inclusion of promoting apoptosis and suppressing proliferation, which indicate the interruption targeting IGF1R ?-strain is prospective for non-small-cell lung carcinoma.

Effect of an Anti－ Keratin Monoclonal Antibody 5G5 on the Proliferation of Cultured Keratinocytes / 中华皮肤科杂志

Objective To investigate the immunological activity and the effect of an anti－ keratin monoclonal antibody 5G5 on the proliferation of serum－ free cultured keratinocytes. Methods The reactivity of 5G5 with cultured keratinocytes was observed by immunohistohemistry technique. The keratins extracted from the cultured keratinocytes was identified by SDS－ PAGE and their reactivities with 5G5 were assessed by immunoblot analysis. MTT was used to study the influences of 5G5 on the proliferation of cultured keratinocytes. Results 5G5 recognized the band of keratin with molecular weight of 50 000 only. Positive staining of 5G5 was found in the cytoplasm of cultured keratinocytes. The strong potential of 5G5 to inhibit the proliferation of cultured keratinocytes was shown by MTT assay with dose－ dependent manner. Conclusions The antibody which specifically recognizes the keratin with molecular weight of 50 000 may be the effective component to inhibit the proliferation of cultured keratinocytes.