Objective To perform genetic analysis in a family line of a pregnant couple with autosomal reces-sive non-syndromic deafness in order to identify its possible genetic etiology and provide prenatal diagnosis.Methods Whole-exome sequencing(WES)was used to analyze the genes of the proband,and Sanger sequencing was used to verify the suspected pathogenic loci.Prenatal genetic diagnosis was performed after amniotic fluid collection at 18 weeks of pregnancy.Results Autosomal recessive deafness type 3 related gene MYO15A c.10419_10423delCAGCT/c.10294_10308delCCTTGCATCCTTGCC compound heterozygous variant was found in the wife.A compound heterozygous variant of autosomal recessive deafness type 77-related gene LOXHD1:c.6388C＞T/ex-on 33-38 del.Maternal MYO15A c.10294_10308del CCTTGCATCCTTGCC heterozygous variant were detected in the husband and paternal LOXHD1 exon 33-38 del heterozygous variant were detected in the fetus.At the same time,the paternal CDH23 c.6693delT heterozygous mutation and the maternal PCDH15 c.5048_5051dupAGAA heterozygous mutation were detected in the fetus.These two heterozygous mutations lead to the possibility of the fe-tus suffering from ID/F Usher syndrome.Conclusion The deafness of the couple is caused by two different deaf gene mutations,and the probability of the fetus having the same deafness as the couple is very low.However,the fetus has a high possibility of having deafness caused by two gene mutations.Therefore,deafness caused by two gene mutations should be paid attention to in the prenatal diagnosis of families with both deaf parents.

OBJECTIVE To investigate the effect and potential mechanism of safflower polysaccharide on apoptosis and autophagy of human hepatocellular carcinoma cells. METHODS Human hepatocellular carcinoma HepG2， SMMC-7721 and Huh-7 cells were selected as subjects， and safflower polysaccharide was used as intervention drug to screen sensitive cells， intervention concentration and intervention time. The sensitive cells were selected as the object and intervened with different concentrations of safflower polysaccharide； the apoptosis， migration， clone formation， morphology and autophagy of human hepatocellular carcinoma cells were observed； the expressions of apoptosis， autophagy and phosphatidyl inositol-3-kinase （PI3K）/protein kinase B （Akt）/mammals rapamycin target protein （mTOR） signaling pathway related protein were detected. RESULTS safflower polysaccharide could inhibit the proliferation of 3 kinds of human hepatocellular carcinoma cells， and the half inhibition concentration of it to SMMC-7721 cells was significantly lower than to other two kinds of cells （P＜0.05）. After 48 h intervened with low， medium and high concentrations of safflower polysaccharide （20， 40， 80 μmol/L）， the apoptosis of SMMC-7721 cells was increased compared with the control group， and cell migration rates at 24 and 48 h （except for safflower polysaccharide low- dose group at 24 h） and clone formation rate at 24 h were significantly decreased compared with the control group （P＜0.05 or P＜ 0.01）. Compared with the control group， cell number in safflower polysaccharide groups was significantly decreased， and autophagy levels were improved to some extent； the relative expressions of cleaved caspase-3， caspase-9， Bax and beclin-1 protein and ratio of LC3Ⅱ/LC3Ⅰ were significantly increased， and the relative protein expressions of Bcl-2， p62， PI3K， mTOR and Akt were significantly decreased （P＜0.05 or P＜0.01）. CONCLUSIONS Safflower polysaccharide could effectively inhibit the proliferation and induce apoptosis and autophagy of human hepatocellular carcinoma SMMC-7721 cells， the mechanism of which may be related to the inhibition of PI3K/Akt/mTOR signaling pathway.

Objective To investigate the influence of supplemental probiotics on the changes of immunity and microecology in asthmatic children. Methods One hundred and seventy?six asthmatic children treated in the Affiliated Hospital of North China University of Science and Technology from October 2015 to October 2016 were selected in the study and were randomly divided into two groups, 88 cases in each group. Patients in the control group were given routine treatment, and the observation group was treated with routine treatment combined with probiotics. The changes in immune index and microecological index before and after the treatment were compared between the two groups. Results After treatment, the observation showed CD3+ was(65. 8±2. 6)％,CD4+was(39. 2±1. 3)％,CD8+ was(24. 5±1. 0)％,CD4+/CD8+ was(1. 6±0. 2),NK cells was(15.2±0.4)％,Th1/ Th2 was(5.7±1.3),interferon γ was(56.3±1.8)ng/L,bifidobacterium was (9. 3±0. 7)lgCFU/g,lactobacillus was(9. 5±0. 6)lgCFU/g,yeast was(6. 6±0. 8)lgCFU/g,compared with those before treatment ((52. 5±1. 7)％,(23. 6±0. 8)％,(19. 7±0. 9)％,(1. 2±0. 1),(12. 8±0. 3)％,(3. 4±0. 7), (44.0±1.5)ng/L,(4.2±1.1)lgCFU/g,(4.9±0.4)lgCFU/g,(3.7±0.4)lgCFU/g),the differences were statistically significant ( t= 5. 533, 9. 957, 5. 436, 6. 332, 4. 875, 9. 764, 5. 727, 15. 143, 12. 387, 10. 837, P<0. 05). After treatment,in the control group,CD3+ was(60. 1±3. 4)％,CD4+ was(30. 7±1. 2)％,CD8+ was (21.9±1.1)％,CD4+/ CD8+ was(1.4±0.3),NK cells was(14.0±0.3)％,Th1/ Th2 was(4.6±0.9), interferon γ was ( 50. 2 ± 1. 4 ) ng/L, bifidobacterium was ( 7. 6 ± 0. 8 ) lgCFU/g, lactobacillus was ( 8. 1 ± 0. 7 ) lgCFU/g, yeast was ( 4. 9 ± 0. 8 ) lgCFU/g, compared with those before treatment ( ( 52. 4 ± 2. 0 )％, ( 23. 8 ±0. 7)％,(19. 8±0. 6)％,(1. 2±0. 2),(12. 7±0. 2)％,(3. 5±1. 1),(44. 1±1. 3)ng/L,(4. 3±0. 9)lgCFU/g, (5.0±0.5)lgCFU/g,(3.8±0.6)lgCFU/g),the differences were statistically significant(t=4.469,5.899, 4. 061,4. 667,4. 023,6. 143,4. 363,10. 674,9. 201,5. 894,P<0. 05) . The above indexes in observation group were higher than those in the control group ( t=3. 948, 3. 162, 4. 187, 4. 428, 3. 857, 5. 391, 4. 202, 5. 236, 4. 728,6. 469,P<0. 05). After treatment,the observation group showed IgE(139. 4±21. 0)was kU/L,IL?4(30. 2 ±1. 7)was ng/L,IL?10 was(6. 3±0. 8)ng/L,escherichia coli was(4. 8±0. 6)lgCFU/g,streptococcus was(6. 1 ±0.9)lgCFU/g,bacillus was(4.6±0.2)lgCFU/g,staphylococcus was(1.9±0.3)lgCFU/g,enterococcus was (5.2±0.4)lgCFU/g,compared with those before treatment((381.2±49.6)kU/L,(59.4±3.5)ng/L,(13.9 ±1.1)ng/L,(7.1±0.5)lgCFU/g,(8.4±0.6)lgCFU/g,(8.0±0.6)lgCFU/g,(4.0±0.8)lgCFU/g,(7.4 ±0. 8)lgCFU/g),while the differences were statictically significant (t=22. 231,12. 667,15. 063,7. 791,6. 770, 10. 392,16. 523,7. 232,P<0. 05). After treatment,in control group showed IgE was (230. 8±31. 7) kU/L,IL?4 was (41. 3±2. 3)ng/L,IL?10 was (9. 8±0. 7)ng/L,escherichia coli was (5. 9±0. 7)lgCFU/g,streptococcus was (7. 2±1. 0)lgCFU/g,bacillus was (6. 4±0. 8)lgCFU/g,staphylococcus was(2. 7±0. 7)lgCFU/g,enterococcus was (6.1±0.6)lgCFU/g,compared with those before treatment((387.9±54.3)kU/L,(59.6±3.4)ng/L, (13. 7±1. 2)ng/L,(7. 0±0. 4)lgCFU/g,(8. 3±0. 5)lgCFU/g,(8. 1±0. 7)lgCFU/g,(4. 1±1. 0)lgCFU/g,(7. 3 ± 0. 7 ) lgCFU/g ) , while there were significant differences ( t= 9. 826, 7. 390, 6. 979, 4. 864, 4. 527, 5. 656、8. 185,4. 967,P<0. 05). The above indexes in observation group were lower than those in the control group(t=9. 618,6. 713,8. 556,5. 290,4. 803,6. 913,7. 215,4. 731,P<0. 05) . The intestinal flora time was ( 5. 6 ± 1) d,hospitalization time was (10. 2 ± 1. 3) d,hospitalization expenses (3527. 1 ± 403. 2) RMB in the observation group,compared with (10. 7±1. 8)d,(14. 6±2. 1)d,(4689. 4±526. 7)RMB in the control group,the differences between the two groups were statistically significant ( t= 12. 107, 7. 314, 6. 295, P<0. 05 ) . Conclusion Probiotic supplement can improve immune status and microecology status in asthmatic children,which is worthy of clinical use.

Objective To explore the clinical value of plasma D -D and blood glucose change in treating patients with bone trauma.Methods Selected 200 patients with bone trauma as the observation group,60 cases of healthy persons as the control group.The plasma D -D and blood glucose of the two groups were detected,and the results were analyzed.Results The plasma D -D and blood glucose of the observation group were more than those of the control group,and it was gradually reduced from first day to third day and fifth day.The plasma D -D of the observation group was (174.7 ±40.2)μg/L,blood glucose was (5.2 ±0.4)mmol/L.The plasma D -D of first day, third day and fifth day of the observation group were (1 452.6 ±303.4)μg/L,(982.2 ±215.2)μg/L,(705.6 ± 152.9)μg/L respectively.The blood glucose of first day,third day and fifth day of the observation group were (14.2 ±2.3)mmol/L,(10.3 ±1.2)mmol/L,(7.6 ±1.9)mmol/L,respectively.The difference was statistically sig-nificant(F =3.49,P <0.05).In the observation group,the more severe injury in patients,the higher plasma D -D and blood glucose,and it was gradually reduced from first day to third day and fifth day,the difference was statistically significant(F =4.83,P <0.05).Conclusion In judging the condition of bone trauma,it is important to measure plasma D -D and blood glucose levels,and this method should be popularized in clinical use.

Objective To explore whether CCL18 is involved in regulating the expression of miRNAs in breast cancer.Methods The expression profile of miRNAs in the breast cancer cell following CCL18 treatment was determined by miRNAs microarray analysis.Then we performed QRT-PCR and Luciferase Reporter Assay to validate the results from the miRNAs microassay.We used transient transfection to change the expression of miR98 and c-myc in breast cancer cells.We then used QRT-PCR and Western blot to analyze the mechanism by which CCL18 downregulates the expression of miR98 in breast cancer cells.Results miRNAs microarray analysis showed that cells treated with CCL18 differentially expressed 20 miRNAs genes compared with those in the control group. Our QRT-PCR and Luciferase Reporter Assay confirmed the result.The mRNA and protein expressions of C-myc and lin28 were increased after CCL18 stimulation in breast cancer cells.Transfection with c-myc siRNAs rescued the increase of lin28 and loss of miR98 expression caused by CCL18 stimulation.Our results also showed that CCL18 could upregulate the expression of N-Ras at post-transcription level.Conclusion CCL18 downregulates the expression of miR98 via N-Ras/c-myc/lin28 pathway.The downregulated miR98 increases the expression of N-Ras after transfection,which further activates c-myc/lin28 pathway and forms a positive feedback loop.

BACKGROUND:Pulmonary infection after kidney transplantation evolves rapidly. There is a high mortality rate in patients with server pulmonary infection. It has the important significance of early diagnosis and treatment of pulmonary infection, but some patients appear to have impaired kidney function because of the adjustment of immunosuppressants. OBJECTIVE:To explore the approaches to applying the immunosuppressants during the treatment of pulmonary infection after kidney transplantation. METHODS:The clinical data of 85 kidney transplantation patients who suffered from pulmonary infection were retrospectively analyzed. There were 43 cases in which the infection occurred within 1-6 months after kidney transplantation, 39 of which within 2-4 months; 7 cases of infection occurring within 6-12 months; 7 cases of infection within 12-24 months; 6 cases of infection within 24-36 months; 22 cases of infection occurring beyond 36 months. The immunosuppressant dose was adjusted based on a per-case basis. As a complement, the smal-dose hormone was used for anti-inflammation. Etiological treatments for resisting infections were also conducted accordingly. Ventilators were utilized for patients with respiratory failures. The body temperature of patients was monitored and controled. Appropriate nutrition support was also provided accordingly. There were 44 cases of decreasing or stopping the use of immunosuppressants during the early period of pulmonary infection; 19 cases of decreasing or stopping the use of immunosuppressants during the treatment of pulmonary infection;5 cases of stopping the use of immunosuppressants during the period of severe pneumonia; 15 cases of gradualy changing the dose of immunosuppressants during the early and progressive period of pneumonia; 2 cases of decreasing the use during the early period of pneumonia and stopping the use during the period of severe pneumonia. The duration of decreasing or stopping the use of immunosuppressants ranged from 3-51 days, with an average of 10.7 days. RESULTS AND CONCLUSION: Among the 85 patients, there were 81 cases cured and 4 cases of death. Among the four death cases, two cases died of acute respiratory failure and two cases died of multiple organ failure. Of the cured 81 cases, acute rejection occurred in 3 cases, while renal alograft dysfunction occurred in 6 cases. Decreasing or temporarily stopping the use of immunosuppressants during the treatment of pulmonary infection caused by the kidney transplantation increases the cure rate and decreases the mortality rate; while timely resuming the usage of immunosuppressants effectively protects the renal graft function, especialy for patients with renal graft dysfunction.

PurposeDiffusion weighted imaging (DWI) can significantly improve the diagnosis of non-enlarged lymph node metastasis. This study aims to evaluate the correlation between the DWI findings and the prognosis, and to identify prognostic factors.Materials and Methods Forty-seven patients with colorectal cancer underwent MRI scan including DWI sequence before surgery. Imaging ifndings were compared with the pathologic results to determine the metastatic lymph nodes (DWI positive) or non-metastatic lymph nodes (DWI negative). Postoperative disease-free survival and overall survival for 5 years of the patients with DWI positive and DWI negative lymph nodes were compared. Correlation between the prognosis and the related factors were investigated including regional DWI-positive lymph nodes, short axis diameter and long axis diameter of the largest DWI-positive lymph node, and number of DWI-positive nodes.Results Of 47 patients,10 (21%) patients had regional DWI-positive lymph nodes showed high signal intensity on diffusion-weighted images. The patients with regional DWI negative lymph nodes had a signiifcant better ifve-year disease-free survival and overall survival (P<0.05). The short axis diameter of the largest DWI-positive lymph node was correlated with distant metastasis (AUC=0.77,P<0.05). The short axis diameter and long axis diameter of the maximum metastatic lymph nodes were correlated with overall survival (AUC=0.84 and 0.75,P<0.05). Five-year disease-free survival and overall survival of the patients with short axis diameter of the largest DWI-positive lymph node ≤9 mm were higher than the patients with lymph node short axis diameter >9 mm (P<0.05). Five-year disease-free survival and overall survival of the patients with long axis diameter of the largest DWI-positive lymph nodes ≤11 mm were higher than the patients with long axis diameter >11 mm (P<0.05). Five-year disease-free survival of the patients with all DWI positive lymph nodes resected was higher than the patient without DWI-positive lymph nodes resected (P<0.05).Conclusion The patients with regional DWI-negative lymph node had a better prognosis. Of the patients with DWI-positive lymph nodes, the patients with smaller lymph nodes have better prognosis than who have larger lymph nodes.

Objective To investigate the effects of sphingosine kinase 1 (SphK1 ) inhibitor N, N-dimethylsphingosine (DMS)combined with 5-fluorouracil (5-FU)on the proliferation and apoptosis of gastric cancer MGC-803 cells and to explore the possible mechanisms involved.Methods MGC-803 cells were cultured in vitro.The effects of DMS and 5-FU on cell proliferation,apoptosis,and cell cycle distribution of MGC-803 cells were detected by MTT assay and flow cytometer (FCM),respectively.The expressions of SphK1 ,TS,DPD,NF-κB p65 and Bcl-2 proteins were detected by Western blot.Results Different concentrations of DMS or 5-FU alone or in combination could obviously inhibit the proliferation of MGC-803 cells in a dose-dependent and time-dependent manners (P<0.05 ).And the proliferation inhibition rate of MGC-803 cells in the combination group was significantly higher than that in the single drug groups (P<0.05).Treatment of MGC-803 cells with DMS did not affect the cell cycle distribution (P>0.05).As compared with the cells without drug treatment,DMS or 5-FU alone could obviously increase the apoptosis rate of MGC-803 cells (P<0.05);the apoptosis rate in the combination group was significantly higher than that in the single-drug groups (P<0.05).The expression levels of SphK1,NF-κB p65 and bcl-2 proteins were down-regulated with the treatment of DMS alone or in combination,whereas those of TS and DPD were not affected.Conclusion DMS can inhibit the proliferation and induce apoptosis of gastric cancer MGC-803 cells in vitro.It shows a good synergetic effect in combination with 5-FU,probably by down-regulating the expressions of SphK1,NF-κB p65 and Bcl-2 proteins.

10.3969/j.issn.2095-4344.2013.25.008

Objective To develop a Gd-based MR probe containing arginine-glyeine-aspartic acid (RGD)motif to reveal integrin avβ3 receptor-expressed tumor.Methods Commercially available HYNICRGD conjugate with co-ligand EDDA was labeled with GdCl3,and the mixture was isolated and purified by solid phase extract(SPE)to get the entire probe Gd-EDDA-HYNIC-RGD.Human HCC cell line BEL-7402 was cultured and the cells harvested and suspended then subcutaneously inoculated into athymic nude mice for tumor growth.In vitro cell binding assay to integrin avβ3 receptor and cell viability experiments were conducted.Then in vivo,imaging of the three arms of xenografts were performed by MR scan with a dedicated animal coil at baseline and time points of 0.30,60,90 mninutes and 24 hour post-intravenous injection(P.i.) via the tail vein.Three arms of nude mice then were sacrificed for histological examination to confirm the imaging results.Results Gd-EDDA-HYNIC-RGD was successfully isolated by SPE and validity was verifled on signal enhancement througll in vitro and in vivo experiments.The T1 relaxation rate of the probe is 3.31 mmol/s:It is well tolerated to living cells when the concentration of the probe is below 0.1 μmol/ml;both BEL-7402 Hunlan Hepatocellular Carcinoma cell Iine and the tumor expressed avβ3receptor;The RGD-iigand was observed specificly binding with avβ3 receptor in vitro;The nude mice model bearing HHCC was well estabhshed.The signal intensity(SI)at the tumor site were 2247.6±39.0 at baseline and 2820.9±35.2 at 90 min p.i.respectively,the SI at 90min increased less than 25%of baseline,which is statistically different(t=-38.031,P＜0.05);while the SI at muscle site were 1824.2±32.8 and 1845.8±27.2 respectively,which is not statistically different(t=-1.424,P＞0.05);The signal to time curve for probe-administrated group is straightforward over time in the span of 0 to 90 minute p.i.while the control arms do not show such tendency.Conclusion Gd-EDDA-HYNIC-RGD has the potential to used as an MR probe detecting integrin avβ3 receptor-expressed tumor.This work may offer possibility of early detection and differentiation of specific tumors.