The megakaryocyte and platelet lineage may be one of the major sites of human cytomegalovirus (HCMV) infection. However, whether HCMV aggravates apoptosis in normal megakaryocytes was not well investigated. Megakaryocytic cell line CHRF-288-11 and HCMV AD 169 strain were co-cultured in this study. PCR was used to detect the direct infection of the cells by HCMV IEA expression. The apoptotic cells were analyzed by morphologic observation, DNA ladder formation, annexin V/PI and PI assay with flow cytometry. The results showed that HCMV significantly inhibited the growth of CHRF cells in three different concentrations of viral infection groups (10(-3), 10(-2), 10(-1)). The viability levels in each infection groups were 77%, 73% and 68% respectively after incubation for 7 days, compared with 98% in the control group. Using annexin V/PI with flow cytometry, it was shown that the percentages of apoptotic cells viral infection in groups (10(-3), 10(-2), 10(-1)) were (21.3 +/- 2.49)%, (25.8 +/- 3.65)% and (31.4 +/- 3.91)% at 7 days after infection, while the control was (3.68 +/- 1.47)%. The apoptotic cells were further confirmed by morphologic observation and DNA ladder formation. Furthermore, PCR detection also showed the direct infection by identification of HCMV IEA expression in CHRF cells. This study suggested that HCMV could directly infect megakaryocytes and aggravated apoptosis in HCMV-infected megakaryocytes.
Apoptosis ; Cell Survival ; Cells, Cultured ; Cytomegalovirus ; pathogenicity ; DNA, Viral ; analysis ; Humans ; Megakaryocytes ; cytology ; virology ; Polymerase Chain Reaction

Objective To evaluate contrast-enhanced fluid-attenuated inversion recovery (FLAIR) imaging in the detection of leptomeningeal lesions.Methods Seventeen patients with a variety of leptomeningeal lesions were analyzed.The MRI protocol included un-enhanced and contrast-enhanced FLAIR images and contrast-enhanced T_1WI,Comparisons between contrast-enhanced FLAIR images and T_1WI and between un-enhanced and contrast-enhanced FLAIR images were made to determine which sequence better depicted the lesions.Results Leptomeningeal lesions showed as either diffusely or locally abnormal hyper-intensity along sulci or cistern on three sequences.Comparison between contrast-enhanced FLAIR and T_1WI showed that only contrast-enhanced FLAIR revealed the abnormalities in 7,both revealed the abnormalities but the former was superior in 2 ,and both were conspicuous in 7. In 1 patient of tuberculous meningitis,diffuse abnormalities of sulci were shown only on contrast-enhanced FLAIR, abnormalities of cisterns were shown on both sequences but the former was superior.Comparison between un- enhanced and contrast-enhanced FLAIR showed that only contrast-enhanced FLAIR revealed the abnormalities in 9,both revealed the abnormalities but the former was superior in 3,and both were conspicuous in 4. In 1 patient of tuberculous meningitis,abnormalities of cisterns were shown only on contrast-enhanced FLAIR,diffuseabnormalities of sulci were shown on both sequences but the former was superior.Conclusions Contrast-enhanced FLAIR images were superior to un-enhanced FLAIR images and contrast-enhanced T_1WI in the detection of leptomeningeal lesions. Contrast-enhanced FLAIR images are helpful and should be considered when findings on un-enhanced FLAIR images and/or contrast-enhanced T,WI are inconclusive.

The study was purposed to investigate the proliferation and the suppression effect of human cytomegalovirus (hCMV) on human promyelocyte cell line HL-60, and to study whether hCMV can induce apoptosis of HL-60 via direct infection in vitro and its mechanism. Promyelocyte cell line HL-60 and hCMV AD169 strain were co-cultured. PCR was used to detect the direct infection of HL-60 cells by hCMV IEA expression. The apoptosis cells were analyzed by morphologic observation, DNA ladder formation, flow cytometry with Annexin V/PI stain. The results indicated that hCMV AD169 suppressed the differentiation and proliferation of HL-60 cells in vitro significantly (P < 0.05). The suppression was dose-dependent. hCMV DNA was successfully detected in HL-60 cells of viral infection groups by PCR. The apoptotic cells were confirmed by morphologic observation and DNA ladder formation. The results of flow cytometry showed that the percentage of apoptotic cells increased along with the increase of hCMV titer and the time after infection. It is concluded that the promyelocyte can be infected directly by hCMV AD169 strain. hCMV AD169 strain inhibited the differentiation and proliferation of promyelocyte. The apoptosis of HL-60 cells can be induced by hCMV infection. With the increase of viral infectious titer and the time after infection, the percentage of apoptotic cells also increase and produce in dose-dependent and time- dependent manner. Induced apoptosis may be one of the mechanisms of granulocytopenia induced by hCMV infection.
Apoptosis ; physiology ; Cell Proliferation ; Coculture Techniques ; methods ; Cytomegalovirus ; physiology ; Cytomegalovirus Infections ; DNA, Viral ; analysis ; Granulocyte Precursor Cells ; virology ; HL-60 Cells ; Humans

OBJECTIVETo investigate the mechanism and the suppression effect of human cytomegalovirus (HCMV) on hematopoietic system.

METHODSSemi-solid culture system was used to observe the effect of HCMV AD169 strain on colony forming unit granulocyte/macrophage (CFU-GM), CFU-erythroid (CFU-E), CFU-multipotent (CFU-Mix) and CFU-megakaryocyte (CFU-MK) growth. The techniques of in situ polymerase chain reaction (IS-PCR) and polymerase chain reaction (PCR) were used to demonstrate the existence of HCMV DNA in the colony cells of cultured CFU-GM, CFU-Mix, CFU-MK and CFU-E, respectively. The immediate early antigen (IEA) mRNA in CFU-MK and late antigen (LA) mRNA in CFU-E were detected by reverse transcriptase-polymerase chain reaction (RT-PCR). HCMV early protein P52 was detected with immunohistochemical technique.

RESULTSHCMV AD169 suppressed the differentiation and proliferation of CFU-GM, CFU-E, CFU-Mix and CFU-MK in vitro significantly (P < 0.05). The suppression was dose-dependent. HCMV DNA was successfully detected in CFU-GM, CFU-Mix, CFU-MK colony cells from viral infection groups by IS-PCR, and was detected in CFU-E by PCR, while it was negative in blank control or mock control groups. CFU-MK colony cells expressed HCMV IEA mRNA with the size of 340 bp in virus infection groups of 10(3) plague forming unit (PFU), 10(4) PFU and 10(5) PFU, respectively. The HCMV LA mRNA was detected by RT-PCR and was 263 bp long in positive control group of HCMV-infected human embryonic fibroblasts. The expression of HCMV LA mRNA in CFU-E was negative. The early protein P52 of HCMV in 10(4) PFU group was also identified by immunohistochemical staining.

CONCLUSIONHCMV AD169 strains inhibited the differentiation and proliferation of CFU-GM, CFU-E, CFU-Mix and CFU-MK by the infection of the hematopoietic progenitors. HCMV might cause the suppression of hematopoiesis by direct infection, which is thought to be one of the reasons of HCMV infection associated with thrombocytopenia, neutropenia and anemia.

Colony-Forming Units Assay ; Cytomegalovirus ; genetics ; DNA, Viral ; genetics ; Erythrocytes ; virology ; Hematopoietic System ; cytology ; virology ; Humans ; Megakaryocytes ; virology ; Multipotent Stem Cells ; virology ; Polymerase Chain Reaction

BACKGROUNDBlood coagulation factor VII (FVII) is physiologically synthesized in the liver and released into the blood. Binding of FVII to tissue factor (TF) is related to the metastatic potential of tumor cells, also a significant risk factor in the development of hepatic metastasis in patients with colorectal cancer (CRC). It has been found that some cancer cells can produce FVII extrahepatically. However, little is known about FVII and CRC. We therefore hypothesized that CRC cells may synthese FVII, leading to tumor invasion and metastasis.

METHODSWe detected the expression of FVII protein in 55 CRC specimens by immunohistochemical staining. The FVII mRNA in 45 of 55 CRC cases, 6 colon cancer cell lines and one hepatoma cell line was measured by real-time reverse transcription-PCR (RT-PCR). Transwell invasion assays were performed to evaluate the changes of cell migration and invasion of LoVo cancer cells in vitro. We further observed the likely effectors regulated by the TF/FVIIa complex Western blotting assay.

RESULTSExtrahepatic synthesis of FVII was detected in the cytoplasm of 32 (58.2%) CRC specimens by immunohistochemistry, but not in normal mucosa. Liver metastasis (P = 0.003) and TNM staging (P = 0.005) were significantly correlated with FVII antigen expression. The positive ratios in stages I, II, III and IV were 33.3%, 40.0%, 52.4% and 87.5%, respectively. The expression of FVII mRNA in CRC with hepatic metastasis was significantly higher than CRC without hepatic metastasis (5.33 ± 2.88 vs. 1.47 ± 0.51, P = 0.03). Ectopic FVIIa induced a slight increase (1.34-fold) in the number of migrating cells, which was inhibited by the specific TF antibody. The formation of TF/FVIIa complex resulted in a marked increase in the expression of matrix metalloproteinases (MMP)-2 (3.5-fold) and MMP-9 (4.7-fold) in a time-dependent and dose-dependent manner.

CONCLUSIONSExtrahepatic synthesis of FVII by CRC cells may promote tumor invasion and metastasis. MMPs, as downstream effectors of TF/FVIIa signaling, facilitate the development of metastasis in colon cancer.

Adult ; Aged ; Aged, 80 and over ; Cell Line, Tumor ; Cell Movement ; Colorectal Neoplasms ; metabolism ; pathology ; Factor VII ; analysis ; biosynthesis ; genetics ; Female ; Humans ; Immunohistochemistry ; Liver Neoplasms ; secondary ; Male ; Matrix Metalloproteinase 2 ; analysis ; Matrix Metalloproteinase 9 ; analysis ; Middle Aged ; Neoplasm Invasiveness ; Neoplasm Staging ; RNA, Messenger ; analysis ; Thromboplastin ; physiology

OBJECTIVETo explore the electrophysiological results and rehabilitation outcome of two prelingually deafened pediatric cochlear implant patients with auditory neuropathy.

METHODSPreoperative audiological evaluation, intra-postoperative electrically evoked auditory brainstem response (EABR) and neural response telemetry (NRT) record for the two cases were conducted in Beijing Tongren Hospital. A one year follow-up was performed. Data collected before and at 6,12-month intervals after implantation were compared with that from control pediatric cochlear implant patients matched for the same duration of implant use as this two cases.

RESULTSThe two children implanted had not had any postoperative medical or cochlear implant device complications. Intraoperative EABR and NRT were elicited in case 1 with unrepeatable waveforms. After 12 months of training, Case 1 had shown significant improvements in sound detection, speech perception abilities and communication skills, which was better than the control group, and the electrophysiological results became normal. Case 2 had also benefited from cochlear implantation, even though no recognizable NRT was found until he returned 12 month after the operation. CONTUSIONS: The desynchronization of auditory path had been changed after the electrical stimulation ongoing 12 months for children with auditory neuropathy. The two children had not had any complications postoperatively, and each child had shown improved listening and communication skills. Cochlear implantation could help patients with auditory neuropathy to improve their communication skill and go back to the main stream.

Child ; Cochlear Implantation ; Cochlear Implants ; Hearing Loss, Sensorineural ; surgery ; Humans ; Infant ; Male ; Retrocochlear Diseases ; surgery ; Treatment Outcome

OBJECTIVETo investigate the sensitive factors which were used in routine audiological tests to find out otitis media with effusion (OME) in newborn infants.

METHODSSubjects of this study were 48 infants, including 31 males and 17 females, who failed in the universal newborn hearing screening. The age ranged from 1.5 to 12 months with the average age of 4.3 months. All subjects accepted temporal bone CT and routine audiological assessments, including air-conduction and bone-conduction auditory brainstem response (ABR), 40 Hz-auditory event related potential (40 Hz-AERP), distortion-product otoacoustic emission (DPOAE), acoustic reflex, tympanometries using 226 Hz and 1000 Hz probe tone. Nine factors were statistically analyzed using Kappa test, Univariate chi(2) test and multivariate condition Logistic stepwise regression analysis, which included the results of acoustic immittance, the air-conduction and bone-conduction ABR thresholds, the difference between air-conduction and bone-conduction ABR thresholds, the latency of ABR wave I, duration between ABR wave I and V, 40 Hz-AERP thresholds, amplitudes and thresholds of DPOAE, and acoustic reflex thresholds (ART).

RESULTSSeventy-seven ears were diagnosed with OME, and 19 ears were normal. CT scan of temporal bone was set as a comparative standard. Kappa test indicated that the results of tympanometry with 1000 Hz probe tone (Kappa = 0.745, P < 0.001), the air-conduction ABR threshold (Kappa = 0.453, P < 0.001), the latency of ABR wave I (Kappa = 0.430, P < 0.001), the threshold of 40 Hz-AERP (Kappa = 0.582, P < 0.001), and DPOAE (Kappa = 0.495, P < 0.001) had agreement with the results of temporal bone CT on evaluating the function of middle ear. Univariate analysis indicated that sensitive factors of middle ear function in newborn infants were tympanometry with 1000 Hz probe tone (P < 0.001), ART (P < 0.001), the air-conduction ABR threshold (P < 0.001), the difference between air-conduction and bone-conduction ABR thresholds (P < 0.001), the latency of ABR wave I (P < 0.001), the threshold of 40 Hz-AERP (P < 0.001) and DPOAE (P < 0.001). And multivariate conditional Logistic stepwise regression model showed that tympanometry with 1000 Hz probe tone (P < 0.001) and 40 Hz-AERP threshold (P = 0.004) can be substituted into Logistic stepwise regression equation.

CONCLUSIONSTympanometry with 1000 Hz probe tone and are sensitive factors to find out OME in newborn infants. The air conduction ABR threshold, ABR wave I latency, 40 Hz-AERP threshold and DPOAE could reflect the middle ear function of newborn infants effectively.

Acoustic Impedance Tests ; methods ; Ear, Middle ; physiopathology ; Evoked Potentials, Auditory ; Female ; Humans ; Infant ; Male ; Otitis Media with Effusion ; diagnostic imaging ; physiopathology ; Sensitivity and Specificity ; Temporal Bone ; diagnostic imaging ; Tomography, X-Ray Computed

OBJECTIVE: To analyze the intellectual landscape and emerging research trends of Chinese medicine (CM) in the management of pediatric asthma through a scientometric study. METHODS: Publications related to CM in the management of pediatric asthma were retrieved from the Web of Science Core Collection using relevant keywords. A scientometric study was performed using CiteSpace and VOSviewer. RESULTS: A total of 1,673 original articles and reviews from 1991 to 2019 were included in the analysis. The amount of annual publications had a gradual increase with time. USA was the major contributor both in country and institution analyses. Based on the co-citation, the published journals were grouped into 4 clusters. Keyword analysis indicated that the main hotspots were: (1) comprehensive management; (2) risk factors, mechanism, and prevalence; (3) prevention and treatment; (4) inflammation; and (5) environmental research. Lastly, we predicted that three emerging trends were quality of life promotion, immune response, and combination therapy. CONCLUSIONS CM research in the management of pediatric asthma will maintain the current trend of steady growth. This scientometric analysis may help scientists to identify the areas of interests and future directions in the field.
Asthma/drug therapy* ; Bibliometrics ; Child ; Humans ; Medicine, Chinese Traditional ; Publications ; Quality of Life