Gene Expression Analysis of Inflammatory Cytokines in Korean Psoriatic Patients

BACKGROUND: Although phenotypic heterogeneity of psoriasis is suggested by the alternate activation of either T-helper (Th)1-related or Th17-related cytokines, little is known about the mRNA levels of inflammatory cytokines. OBJECTIVE: To investigate whether there is differential expression of Th1-related and Th17-related inflammatory cytokine genes 1) between psoriatic patients and healthy controls, and 2) between patients with different psoriasis phenotypes. METHODS: Twenty-five patients with psoriasis (10 with guttate psoriasis and 15 with plaque psoriasis) and 5 healthy volunteers were enrolled in this study. The mRNA levels of circulating cytokines (interleukin [IL]-2, IL-12p40, interferon-γ, IL-17A, IL-22, and IL-23R) were measured by real-time reverse transcription polymerase chain reaction. RESULTS: The comparison between psoriatic and healthy control samples revealed that IL-12p40, IL-17A, and IL-22 mRNA levels were significantly higher (approximately 4∼6 folds) in the patients with psoriasis. The mRNA levels of these six cytokines in the blood did not differ between the guttate and plaque psoriasis groups. CONCLUSION: We found that the mRNA levels of blood inflammatory cytokines (IL-12p40, IL-17A, and IL-22) were significantly elevated in patients with psoriasis compared to the levels in healthy controls, but they did not significantly differ between patients with guttate and plaque type psoriasis.

Recombinant AAV Vector with MITF-M Promoter for Melanoma Gene Therapy

We have developed the recombinant adeno-associated virus (AAV) carrying the EGFP gene under the control of the microphtalmia-associated transcription factor-M (MITF-M) promoter region for melanoma-specific expression. MITF-M distal enhancer (MDE) region enhances the specific expression of the reporter gene specifically in cultured melanoma cells. Expression of EGFP protein was very high in AAV-CMV-EGFP infected cells but relatively low in cells infected with AAV-Mitf(Enh/Pro)-EGFP. After an in vitro infection by a recombinant AAV carrying the EGFP gene under the control of human MITF-M promoter, the reporter gene was expressed in MITF-M producing melanoma cell lines (SK-28 and G361), but not in MITF-M non-producing cell lines (HaCat). These results suggest that the utilization of the MITF-M promoter in a recombinant AAV vector could provide benefits in gene therapy applications.

Inactivation of 2009 Pandemic Human Influenza A Virus H1N1 by Photocatalyst Under UV Irradiation

A large-scale pandemic by human influenza virus H1N1 in 2009 caused severe health, social, and economic impacts. In this study, a photocatalyst technology based on TiO2, was evaluated for inactivation of a human influenza virus H1N1 isolated from a patient. The virus titer was reduced by 103.16-fold within 24 h and more than 104.31-fold inactivation within 48 h and 72 h. These results suggest that the tested photocatalyst technology based on TiO2 can be used for reduction of influenza A virus adherence to other surfaces with Hizen-s inside diverse buildings, enabling effective control of its indirect contact infection. The photocatalyst is expected also to reduce level of the aerosol transmission of the virus.

Investigation of Murine Norovirus Replication in RAW264.7 Cells by Strand-specific RT-PCR

Murine norovirus (MNV) is a non-enveloped virus with a positive-sense RNA genome and causes lethal infection in mice. MNV has been used as a model virus for human norovirus (NV) whose in vitro cell culture system has not been available to date since MNV and NV are genetically related. In this study, the genome replication of MNV was investigated using strand-specific RT-PCR in RAW264.7 cells. Reverse transcription (RT) using a sense primer followed by PCR showed that negative-sense RNAs were first detected in RAW264.7 cells between 6 and 9 [3 and 6] hours post infection (h.p.i.). However, these negative-sense RNAs were not detected when cells were treated with a translation inhibitor cycloheximide. Then, RT with an antisense primer followed by PCR was performed to detect positive-sense RNAs. RT-PCR results revealed that the amount of positive-sense RNAs began to increase from 9 [6] h.p.i., indicating the accumulation of the newly synthesized (+)RNA genome. Furthermore, cycloheximide abrogated the increase of newly made RNAs during MNV infection. In conclusion, strand-specific RT-PCR using a sense or antisense primer, in combination with cycloheximide treatment, enabled us to detect positive-sense and negative-sense RNAs selectively and provided a useful tool to understand the replication cycle of MNV.

Protein Expression of the Human Norovirus Capsid Gene using the Baculovirus Expression System

Human norovirus (HuNoV) is the major etiological agent of nonbacterial gastroenteritis worldwide. However, due to the absence of a rapid and sensitive diagnostic system, detection and monitoring have been limited. The HuNoV genome is composed of three open reading frames (ORFs). And major capsid protein, ORF2, is designated as a viral protein 1 (VP1). In this study, the baculovirus expression system was used for expression of the HuNoV capsid protein, VP1. Recombinant baculoviruses can be used as potent tools in HuNoV studies.

Antibody Production of Baculovirus-expressed VP6 from Porcine Group C Rotavirus

The emerging pathogen, group C rotavirus (RVC) has been reported to cause acute diarrhea. But there was the limitation on the detection and monitoring for the absence of rapid sensitive diagnosis system. For the molecular biology study and diagnostic system development, we could detect porcine RVC by reverse transcriptase PCR (RT-PCR) analyses from 60 diarrheal disease porcine stool samples. VP6 full length RT-PCR product (CA-2 RVC, 1352 bp) was cloned and compared the nucleotide and deduced amino acid sequences with those of previously reported other porcine, human, and bovine rotavirus group A, B and C strains. Analyses data showed >82% homology on the nucleotide sequences and >90% homology on the deduced amino acid sequences with other RVCs. Recombinant baculovirus was prepared with cloned PCR product corresponding to VP6 coding sequence (CDS) (position 22~1206) into BaculoDirect(TM) C-term linear DNA, and used for the transfection of insect cells. The polyclonal antibody was produced from mice with purified recombinant VP6 and confirmed with western blot. Both of VP6 antigen and antibody, are useful for the development of rapid diagnostic system against RVC.

Antiviral Activity of Lactobacillus spp. and Polysaccharide

Lactobacillus species have been widely used in both human and animals to prevent or treat gastrointestinal disorders. Recently, it was reported that Lactobacillus spp. inhibited infections by respiratory and gastroenteric viruses; however, its mechanism is not clear. Lactobacillus spp. play direct and indirect roles in the inhibitory effects of viral replication. 1) In vitro study: Highest protection effects were showed with the known probiotics L. rhamnosus GG (LGG) and L. casei Shirota against both rotavirus (RV) and transmissible gastroenteritis virus (TGEV). 2) In vivo study: L. acidophilus had significant immunopotentiating effects, and was therefore recommended for use as a safe oral adjuvant for rotavirus vaccines in pigs. Oral administration of lactobacilli, such as LGG and L. gasseri, might protect a host animal from influenza virus (IFV) infection. Polysaccharides are regarded to be potentially useful and biologically active as an ingredient for pharmaceutical uses due to a variety of biological activities. Especially, sulphated polysaccharides exhibit broad-spectrum antiviral activity against enveloped viruses in vitro. With respect to human immunodeficiency virus type 1 (HIV-1), its in vitro antiviral activity is, specifically, the inhibition of virus-cell attachment, the first step in the infection process. Recently, it was reported that sulphated polysaccharides exhibited antiviral activity against HBV, HCMV, HSV and IFV. In conclusion, Lactobacillus spp. and polysaccharides with antiviral activity against diverse viruses are potential candidates as ingredients for probiotics and medicine candidate for the prevention and treatment of viral infections in animals and humans.

Production and Quality Control of Adenoviral Vectors for Clinical Trials

The importance of recombinant adenoviral vectors for the development of gene therapy and prophylactic and therapeutic vaccines has led to efforts for process development of large scale production of clinically safe adenoviral vectors. First of all, cell lines producing replication incompetent adenoviral vectors required for clinical application have been developed and the concept of banking and characterization of cell lines and adenoviral vectors has been established. In order to meet the need of amount of adenoviral vectors for clinical trials, various large scale suspension culture methods using serum-free media have been developed along with development of large scale purification methods using chromatography instead of cesium chloride method. In addition, methods for the quality control of adenoviral vectors have been established and applied for the clinical lots.

Purification of Protein Expressed from Three Different Regions of Norovirus (NoV)

Norovirus (NoV), which belongs to the family Caliciviridae, is one of the major causes of nonbacterial acute gastroenteritis in the world. In this study, we purified proteins from the epitope region of norovirus for development of the rapid diagnosis system using polyclonal antibodies. As antigens, parts of the ORF (open reading frame) 2, ORF2-P domain, ORF2-Epi, and ORF3 regions were selected and their expressions were induced. The antigenicity of the purified proteins was identified by Western blotting. Each of the purified proteins was injected into mice for the production of novel antibodies and after 3 months of immunization, sera from the mice were obtained. The polyclonal antibody titer was tested by enzyme-linked immunosorbent assay (ELISA) and antibody against ORF2-Epi showed the highest titer. Those polyclonal antibodies can be used in further immunoassay for the rapid detection of NoVs from food and clinical specimens.

A Experimental Model of Bladder Cancer Using Nonviral Vector Expressing Uroplakin II Promoter and SV40 T Antigen / 대한비뇨기과학회지

PURPOSE: The goal of this study was to constructing a nonviral vector, expressing the chimeric gene of the SV40 T antigen and mouse uroplakin II promoter (UPII promotor), which was uniquely expressed in the urothelium, to aid in the treatment of bladder cancer by the creation of a tumor that will express itself in the bladder only, but that will have no effect on the other urothelium. MATERIALS AND METHODS: 36 female C3H/He mice, weighing 20-25grams, were used in this study. A UPII-GFP-liposome complex was installed into the bladder, with Enhanced Green Fluorescent Protein (EGFP), expressing the bladder mucosa, and analyzed via fluorescent microscopy. A UPII- SV40T-liposome complex was then administered into the bladders of the mice, and the bladder and ureter examined, grossly and microscopically, at 1, 2, 3 and 4 weeks, to find transitional cell carcinomas specific to the bladder, the degree of bladder cancer development, and whether the development was from superficial to deep tumors, as well as tumor metastasis. RESULTS: The expression of EGFP was found in all four mice after 2 days. No development of tumors was evident in any mice. However, of the 6 mice sacrificed 28 days after bladder instillation, urothelial dysplasia was evident in 4. There was no evidence of transient cell carcinomas in the ureter or renal pelvis in any of the mice, or of distant metastasis during the term of the study. CONCLUSIONS: This model of bladder cancer seems to take longer than other models for cancer formation as the carcinogen affects the DNA of urothelial cell for the formation of bladder cancer. However, our bladder cancer model was better than others due to its similarity for the processes of normal bladder cancer formation.

The Temperature Differences among the Three Urethral Portions (Distal, Middle & Proximal) and Bladder in Incontinent Women

PURPOSE: We tried to find out an adequate sol-gel transition temperature of female urethra for the injection of thermosensitive polymer in incontinent patients. We measured the temperatures of three portions of female urethra and bladder. MATERIALS AND METHODS: Total of 53 female incontinent patients participated, excluding those with any kind of infection which could lead to an elevation of body temperature. The basal body temperatures were checked at the axilla, tympanic membrane and mouth. Temperatures of the proximal(U1), middle(U2), distal(U3) urethra and bladder(B) were measured by a digital thermometer under a lithotomy position. We divided our patients into 3 groups which were patients in follicular phase(F), luteal phase(L) and menopause(M). The temperature difference between the 4 portions of the urethra(D1; between U1 and U2, D2; between U2 and U3, D3: between U3 and B), was also analyzed. Statistics was done by the ANOVA of repeated measures, one-way ANOVA and Pearson correlation coefficient. RESULTS: The mean age of the patients was 48.1+/-10.7 years. The mean temperature of B, U1, U2, and U3 groups were 37.1+/-0.25 degreesC, 37.0+/-0.25 degreesC, 36.9+/-0.24 degreesC, and 36.7+/-0.25 degreesC. The mean temperature difference of D1, D2, and D3 were 0.2471+/-0.089 degreesC, 0.079+/-0.066 degreesC and 0.066+/-0.058 degreesC. The Pearson correlation coefficient of D1, D2 and D3 were 0.938, 0.965 and 0.970. This showed there was a constant temperature increase from distal urethra to bladder step by step. The number of patients in F, L and M groups were 25(47.2%), 10(18.9%) and 18(33.9%). There was no significant urethral temperature difference at each point(U1, U2, U3 and B) among these three groups. CONCLUSION: There was a constant temperature increase from distal urethra to bladder step by step. This is a baseline study for female urethra for future clinical study. We suggest that our data can be used as deciding the sol-gel transition temperature for thermosensitive polymer injection into incontinent female urethra.

The Isolation and Characterization of Muscle Derived Stem Cells from Gastrocnemius Muscle of Rats Using the Modified Preplate Method / 대한비뇨기과학회지

PURPOSE: This study attempted to characterize the muscle derived stem cells isolated from the primary cultured skeletal muscle of the rat gastrocnemius muscle; in addition, we modified the preplate method and then compared this to the original preplate method. MATERIALS AND METHODS: The hind limbs (gastrocnemius muscles) were removed from a 3-6 week olds SD-rat and the bone was dissected away. The muscle mass was finely minced and chopped using razor blades. In an original preplate method, the cells were dissociated using a triple enzyme mixture (collagenase XI, dipase and trypsin) for 1 hour at 37degreesC. The muscle cell extract was preplated on culture flasks as described by Dr. Qu (Qu et al., 1998). The pp1-pp4 cells were referred to as the early plate (EP) cells, and the pp5-pp6 cells were referred to as the late plate (LP) cells. When we modified the preplate method, the pp1-pp2 cells were called the early plate (EP) cells and the pp3-pp4 cells were called to late plate (LP) cells. The phenotypical characteristics of EP and LP cells were compared by immunostaining and FACS. RESULTS: In the original preplate methods, the early plate (EP) cells were mixed with myogenic cells (mostly fibroblasts, 90% desmin + cells). Yet in the modified preplate method, the muscle derived stem cells were determined to be CD34 (+ or -), CD45- and desmin- cells by immunohistochemical staining and FACS. CONCLUSIONS: In original methods, the LP cells exhibited stem cell properties (CD34+, less than 30%), and they were not from a hematogeous origin (CD45-), but rather, they were from a myogenic origin (desmin+). Yet in the modified preplate method, we purified the LP cells much earlier than the original method. The LP cells displayed CD34+(more than 50%), and CD45-; thus, we isolated more primitive (desmin-) cells.

The Isolation and Characterization of Muscle Derived Stem Cells from Gastrocnemius Muscle of Rats Using the Modified Preplate Method / 대한비뇨기과학회지

PURPOSE: This study attempted to characterize the muscle derived stem cells isolated from the primary cultured skeletal muscle of the rat gastrocnemius muscle; in addition, we modified the preplate method and then compared this to the original preplate method. MATERIALS AND METHODS: The hind limbs (gastrocnemius muscles) were removed from a 3-6 week olds SD-rat and the bone was dissected away. The muscle mass was finely minced and chopped using razor blades. In an original preplate method, the cells were dissociated using a triple enzyme mixture (collagenase XI, dipase and trypsin) for 1 hour at 37degreesC. The muscle cell extract was preplated on culture flasks as described by Dr. Qu (Qu et al., 1998). The pp1-pp4 cells were referred to as the early plate (EP) cells, and the pp5-pp6 cells were referred to as the late plate (LP) cells. When we modified the preplate method, the pp1-pp2 cells were called the early plate (EP) cells and the pp3-pp4 cells were called to late plate (LP) cells. The phenotypical characteristics of EP and LP cells were compared by immunostaining and FACS. RESULTS: In the original preplate methods, the early plate (EP) cells were mixed with myogenic cells (mostly fibroblasts, 90% desmin + cells). Yet in the modified preplate method, the muscle derived stem cells were determined to be CD34 (+ or -), CD45- and desmin- cells by immunohistochemical staining and FACS. CONCLUSIONS: In original methods, the LP cells exhibited stem cell properties (CD34+, less than 30%), and they were not from a hematogeous origin (CD45-), but rather, they were from a myogenic origin (desmin+). Yet in the modified preplate method, we purified the LP cells much earlier than the original method. The LP cells displayed CD34+(more than 50%), and CD45-; thus, we isolated more primitive (desmin-) cells.

The Effect of Recombinant Baculovirus (BacG-CMV-P53) Mediated Gene Therapy in the HT-1376 Bladder Cancer Cell Line / 대한비뇨기과학회지

PURPOSE: In preliminary studies, it was found that mammalian cells can be infected by recombinant baculovirus in vitro. Therefore, the potential use of recombinant baculovirus(BacG-cytonegalovirus(CMV)-P53) for the bladder gene therapy was investigated. MATERIALS AND METHODS: The recombinant Baculovirus(BV) pseudotyped was developed with the vesicular stomatitis virus(VSV) G protein. The presence of the VSV-G protein in purified BV preparations was confirmed by Western blotting analysis. The bladder cancer cells of human(HT-1376) were infected with various multiplicity of infection(MOI) of the BV, and the percentage of apoptotic cells determined by methyl thiazolyl tetrazolium(MTT) assay. RESULTS: The suppression effect of the recombinant BV with a P53 insertion in human bladder cancer cells(HT-1376) increased as the MOI of the recombinant BV increased; 100% cell survival in the group with PBS, and 81.6+/-4.3, 52.0+/-5.6 and 39.8+/-3.7% at 1, 10 and 100 MOI, respectively (p<0.05). CONCLUSIONS: Significant growth suppression was observed following infection with BacG-CMV-P53 in a human bladder cancer cell line. This observation suggests that BacG-CMV-P53 may be a potentially effective agent to prevent recurrence for P53 mutated bladder cancer. Bladder gene therapy using recombinant baculovirus could be a safe and effective treatment of bladder cancer.

The Effect of Recombinant Baculovirus (BacG-CMV-P53) Mediated Gene Therapy in the HT-1376 Bladder Cancer Cell Line / 대한비뇨기과학회지

PURPOSE: In preliminary studies, it was found that mammalian cells can be infected by recombinant baculovirus in vitro. Therefore, the potential use of recombinant baculovirus(BacG-cytonegalovirus(CMV)-P53) for the bladder gene therapy was investigated. MATERIALS AND METHODS: The recombinant Baculovirus(BV) pseudotyped was developed with the vesicular stomatitis virus(VSV) G protein. The presence of the VSV-G protein in purified BV preparations was confirmed by Western blotting analysis. The bladder cancer cells of human(HT-1376) were infected with various multiplicity of infection(MOI) of the BV, and the percentage of apoptotic cells determined by methyl thiazolyl tetrazolium(MTT) assay. RESULTS: The suppression effect of the recombinant BV with a P53 insertion in human bladder cancer cells(HT-1376) increased as the MOI of the recombinant BV increased; 100% cell survival in the group with PBS, and 81.6+/-4.3, 52.0+/-5.6 and 39.8+/-3.7% at 1, 10 and 100 MOI, respectively (p<0.05). CONCLUSIONS: Significant growth suppression was observed following infection with BacG-CMV-P53 in a human bladder cancer cell line. This observation suggests that BacG-CMV-P53 may be a potentially effective agent to prevent recurrence for P53 mutated bladder cancer. Bladder gene therapy using recombinant baculovirus could be a safe and effective treatment of bladder cancer.

Seroepidemiological Survey of Hantaan Virus Infection in Sera of Patients at the Kangnam St. Mary Hospital in Korea from 1998 to 2002

Sera of patients visited at the Kangnam St. Mary Hospital in Seoul were collected randomly at the Department of Clinicopathology from January, 1998 to December, 2002. Specimens were collected two twice a month, in a 15-day interval, and 100 specimens were collected at a time. Specimens test in duplicate, and/or displaying antinuclear antibody reaction were excluded from the seroepidemiological analyses. Detection of antibodies to Hantaan virus, an etiologic agent of hemorrhagic fever with renal syndrome (HFRS), was done by indirect immunofluorescent antibody (IFA) technique. Out of 11,361 sera tested, 445 cases (3.9%) showed specific antibody to Hantaan virus. Sexual difference was not noted. Annual incidence of HFRS cases showed a 3 year-periodicity. In the monthly incidence analysis, two peaks of incidence were appeared in the male cases, the first peak in March and the second in August. Female cases showed a single peak in October. The age distribution showed that 64.9% of the sero-positive cases were from 40 to 69 years of age. Peak age-group was in the 6th decade. Each decade of age-group showed diverse patterns of annual and monthly incidences. These results suggest the incidence of HFRS shows a periodicity and a unique pattern in each age group.

Associations of HLA Alleles with Chronic Infection and Prophylaxis in Vertical Transmission of Hepatitis B Virus

Perinatal transmission and infection of hepatitis B virus (HBV) in early childhood were observed in the offsprings of hepatitis B surface antigen (HBsAg)-positive mothers who had been vaccinated against HBV immediately after giving birth. This prophylaxis failure of perinatal HBV infection is likely due to the interplay of the virus and host immune response. To investigate whether the HLA polymorphism affected the outcome of the perinatal prophylaxis, HLA class I (HLA-A, B and Cw) and class II (HLA-DRB1, DQA1, DQB1 and DPB1) were typed using serology, PCR-SSOP (polymerase chain reaction-sequence specific oligonucleotide probe), and PCR-ARMS (amplification refractory modification system) methods in 22 HBeAg-positive mothers and their 10 prophylaxis-succeeded and 12 prophylaxis- failed children. The HLA types of the mothers and their children were compared with 198 HBsAg-negative healthy controls in a Korean population. HLA-B35 (relative risk=4.2, p<0.01), B51 (relative risk=3.2, p<0.02), DRB1*07 (relative risk=3.8, p<0.03), and DQA1*02 (relative risk=3.8, p<0.03) alleles were more frequent in HBeAg-positive mothers than in the controls. Also, HLA-DRB1*13 (relative risk=0.1, p<0.02) and DPB1*0401 (relative risk=0.1, p<0.02) alleles were less frequent in HBeAg-positive mothers. However, HLA alleles did not affect the outcome of the perinatal prophylaxis against HBV. These results suggest that the reported influences of some HLA alleles on the natural chronic HBV infections may not operate in the HBV infections in children received perinatal prophylaxis.

Characteristics of Respiratory Tract Infection in the Hematopoietic Stem Cell Transplantation Population / 감염

PURPOSES: The respiratory tract infection is one of the most prevalent and serious complications following hematopoietic stem cell transplantation (HSCT). Reports not only for the respiratory tract infection but, unlikely for bacteria or fungi, for the infections caused by the respiratory viruses have been rarely reported in Korea. During the winter of 2000~2001, authors wanted to know the prevalence rate of the respiratory tract infection and the kinds of causative microorganisms, especially the community respiratory viruses (CRV). Based on these data, we attempted to evaluate the clinical courses and prognosis of the patients. METHODS: From October 2000 to February 2001, specimens were collected from the patients who visited Catholic hemopoietic stem cell transplantation center, showing symptoms and signs of respiratory tract infection after HSCT. Standard methods have been applied to isolate and identify bacterial and fungal species. Measles was diagnosed based on the typical symptoms, rash, fever, and Koplik spot. For the four different CRV (adenovirus, RSV, influenza virus, parainfluenza virus), multiplex PCR and conventional culture method were used for the identification. RESULTS: Eighty-four specimens were collected from 66 patients for 4 month period. Average age of patients was 35+/-8 years. Sixty patients (90%) were received allogeneic HSCT. Sample collection was performed between 10 and 3,740 days (average 370 days, median 215 days) after HSCT. Forty-seven patients (71.2%) have been received immunosuppressants at the time of respiratory tract infection. Forty patients (60.6 %) were suffered lower respiratory tract infection and forty-four patients (66.7%) had community-acquired infection. Sixty microorganisms were identified from 45 patients out of total 66 patients. Identified microoganisms were bacteria accounting for 2 cases (3.4%), fungi for 11 (18.3%), tuberculosis for 5 (8.3%), and viruses for 42 (70.0%). Among viruses, 16 cases were measles (39%), 14 adenovirus (33%), 9 cytomegalovirus (21%), 2 parainfluenza virus (5%), 1 was influenza virus (2%). However, no RSV was identified. Most of patients showed good prognosis without any complications. Ten (15.2%) out of total 66 patients were expired. The direct cause of death for all 8 among 10 patients was pneumonia. CONCLUSION: Of the respiratory tract infection fol-lowing HSCT, most common causative microorganisms were viruses - measles, adenovirus in order. No case of RSV infection was found. No epidemic must be occurred by influenza virus because only 1 case was found. Fourteen patients were infected by more than one microorganisms. Overall mortality rate was 15.2%. This study is still undergoing and once accumulated data for more than 1 year, it might be possible to work out a strategies of treatment and prevention for respiratory tract infections. We also expect that these data might be able to provide the basis of efficient infection control in HSCT unit.

Frequency and Clinical Manifestations of Human Herpesvirus-6 Infection in Hematopoietic Stem Cell Transplant Recipients / 감염

BACKGROUND: Human herpesvirus-6 (HHV-6) is recently known as a major pathogen associated with various diseases in hematopoietic stem cell transplant (HSCT) recipients. We prospectively evaluated the frequency and clinical manifestations of HHV-6 infection in HSCT recipient in a single HSCT center in Korea. METHODS: Serum and peripheral blood mononuclear cells (PBMC) were weekly obtained from 1 week before HSCT to 4 weeks after HSCT. Three months' and six months' samples were obtained in some cases. HHV-6 was detected by nested polymerase chain reaction. RESULTS: Two hundred and seventy-eight samples from 54 HSCT recipients were collected from February to November, 1999. HHV-6 was detected in 32 out of 54 recipients (59.3%) at least once during study period in their PBMC or serum. HHV-6 DNA positive rate of PBMC and serum samples were 38.1% and 4.3 % respectively. HHV-6 DNAemia (HHV-6 DNA positive in serum) was detected and peaked at 2 weeks after HSCT and continued to 4 weeks. HHV-6 DNA in peripheral blood was not associated with unexplained fever, acute graft-versus-host disease, engraftment delay, or cytomegalovirus infection in this study. CONCLUSION: Reactivation and development of DNAemia of HHV-6 certainly occurred after HSCT, but the clinical manifestations and association with other diseases were unclear in this study. The large-scaled, nation-wide detail studies about the prevalence and characteristics of HHV-6 in general population and patients of specific disease entities must be considered.

Pseudotype HIV-1 Particles Carrying CD4

A defective HIV-1 helper virus DNA, pHyPC, was assembled by deleting the RNA packaging signal, env, nef and the 3'LTR sequences. HIV-1 like virus particles that carry the HIV-1 receptor, CD4 were generated by coexpression of pHyPC and plasmid DNAs encoding different chimeric CD4 proteins. The CD4 particles, sharing the CD4 ectodomain, precisely fused to different membrane anchors. CD4(+) particles specifically bound to HIV-1 Env expressing cells, but any signs of infection into these cells were not detected. Binding was only partially blocked by either polyclonal anti-CD4 antibodies or by high concentrations of soluble CD4. Suprisingly, CD4(+) particles also adsorbed to HeLa, CHO, NIH3T3 and COS-7 cells in the absence of HIV-1 Env expression. Adsorption was comparable in strength and speed to the highly specific CD4-Env interaction. CD4(-) particles exhibited only background levels of binding. Cell binding was CD4- dependent, but it was independent of the cell type from which the CD4(+) particles originated. Interestingly, CD4-dependent/Env-independent binding was only found when CD4 was present on virus particles. This suggests that the micro-environment of CD4 on virus particles uniquely expose this new cell binding activity. Its high affinity could explain in part why infection of Env(+) cells by CD4(+) particles was not detected. Further experiments will be required to evlauate whether this strong membrane interaction could represent one step in the multiple-step viral entry process.