Cloning of ABO PCR Products Including Exon 2 to Exon 7 Using Seamless Cloning Technique / 대한수혈학회지

Background@#Sequencing and allele separation from exon 2 to exon 7 in the ABOgene can be required to determine the genotype. This study used the seamless cloning technique to clone approximately 7.3 kb containing a portion of exon 2 to exon 7 of the ABO gene. @*Methods@#The previously confirmed DNA sample of the ABO*B.01/O.01 genotype was used for seamless cloning using the Overlap Cloner DNA Cloning Kit (ElpisBio). Cloning was performed three times using the same cloning protocol. The plasmids obtained by cloning were also confirmed by sequencing. @*Results@#Ninety-two colonies (average of 30 colonies per culture plate) were obtained. Of the fifty colonies, 48 colonies were properly amplified. Plasmid sequencing confirmed that 31 heterozygous alleles were well separated in accordance with predictions through the BGMUT database. @*Conclusion@#The data suggest that the seamless cloning technique is a reliable technique that can be used to separate long-range alleles, such as the ABO gene.

Cloning of ABO PCR Products Including Exon 2 to Exon 7 Using Seamless Cloning Technique / 대한수혈학회지

Background@#Sequencing and allele separation from exon 2 to exon 7 in the ABOgene can be required to determine the genotype. This study used the seamless cloning technique to clone approximately 7.3 kb containing a portion of exon 2 to exon 7 of the ABO gene. @*Methods@#The previously confirmed DNA sample of the ABO*B.01/O.01 genotype was used for seamless cloning using the Overlap Cloner DNA Cloning Kit (ElpisBio). Cloning was performed three times using the same cloning protocol. The plasmids obtained by cloning were also confirmed by sequencing. @*Results@#Ninety-two colonies (average of 30 colonies per culture plate) were obtained. Of the fifty colonies, 48 colonies were properly amplified. Plasmid sequencing confirmed that 31 heterozygous alleles were well separated in accordance with predictions through the BGMUT database. @*Conclusion@#The data suggest that the seamless cloning technique is a reliable technique that can be used to separate long-range alleles, such as the ABO gene.

Inhibition of ERK1/2 by silymarin in mouse mesangial cells

The present study aimed to show that pro-inflammatory cytokines [tumor necrosis factor (TNF)-α, interferon (IFN)-γ, and interleukin (IL)-1β] synergistically induce the production of nitric oxide (NO) production in mouse mesangial cells, which play an important role in inflammatory glomerular injury. We also found that co-treatment with cytokines at low doses (TNF-α; 5 ng/ml, IFN-γ; 5 ng/ml, and IL-1β; 1.25 U/ml) synergistically induced NO production, whereas treatment with each cytokine alone did not increase NO production at doses up to 100 ng/ml or 50 U/ml. Silymarin, a polyphenolic flavonoid isolated from milk thistle (Silybum marianum), attenuates cytokine mixture (TNF-α, IFN-γ, and IL-1β)-induced NO production. Western blot and RT-PCR analyses showed that silymarin inhibits inducible nitric oxide synthase (iNOS) expression in a dose-dependent manner. Silymarin also inhibited extracellular signal-regulated protein kinase-1 and -2 (ERK1/2) phosphorylation. Collectively, we have demonstrated that silymarin inhibits NO production in mouse mesangial cells, and may act as a useful anti-inflammatory agent.

Influence of Fimasartan (a Novel AT1 Receptor Blocker) on Catecholamine Release in the Adrenal Medulla of Spontaneously Hypertensive Rats

The aim of this study was to determine whether fimasartan, a newly developed AT1 receptor blocker, can affect the CA release in the isolated perfused model of the adrenal medulla of spontaneously hypertensive rats (SHRs). Fimasartan (5~50 microM) perfused into an adrenal vein for 90 min produced dose- and time-dependently inhibited the CA secretory responses evoked by ACh (5.32 mM), high K+ (56 mM, a direct membrane depolarizer), DMPP (100 microM) and McN-A-343 (100 microM). Fimasartan failed to affect basal CA output. Furthermore, in adrenal glands loaded with fimasartan (15 microM), the CA secretory responses evoked by Bay-K-8644 (10 microM, an activator of L-type Ca2+ channels), cyclopiazonic acid (10 microM, an inhibitor of cytoplasmic Ca(2+)-ATPase), and veratridine (100 microM, an activator of Na+ channels) as well as by angiotensin II (Ang II, 100 nM), were markedly inhibited. In simultaneous presence of fimasartan (15 microM) and L-NAME (30 microM, an inhibitor of NO synthase), the CA secretory responses evoked by ACh, high K+, DMPP, Ang II, Bay-K-8644, and veratridine was not affected in comparison of data obtained from treatment with fimasartan (15 microM) alone. Also there was no difference in NO release between before and after treatment with fimasartan (15 microM). Collectively, these experimental results suggest that fimasartan inhibits the CA secretion evoked by Ang II, and cholinergic stimulation (both nicotininc and muscarinic receptors) as well as by membrane depolarization from the rat adrenal medulla. It seems that this inhibitory effect of fimasartan may be mediated by blocking the influx of both Na+ and Ca2+ through their ion channels into the rat adrenomedullary chromaffin cells as well as by inhibiting the Ca2+ release from the cytoplasmic calcium store, which is relevant to AT1 receptor blockade without NO release.

Synergistic Induction of iNOS by IFN-gamma and Glycoprotein Isolated from Dioscorea batatas

Dioscorea species continue to be used in traditional Chinese medicine, and represent a major source of steroid precursors for conventional medicine. In the previous study, We isolated glycoprotein (GDB) from Dioscorea batatas, characterized, and demonstrated immunostimulating activity in C57BL/6 mice. The aim of this study was to investigate the mechanism whereby GDB activates macrophages. Macrophages activation by GDB was investigated by analyzing the effects of GDB on nitric oxide (NO) production, iNOS expression, mitogen activated protein kinase (MAPK) phosphorylation, and transcription factor activation. In the presence of IFN-gamma, GDB strongly stimulated macrophages to express iNOS and produce NO. Furthermore, the activation of p38 was synergistically induced by GDB plus IFN-gamma , but SB203580 (a p38 inhibitor) inhibited GDB plus IFN-gamma-induced p38 activation. This study indicates that GDB is an important activator of macrophages. Furthermore, due to the critical role that macrophage activation plays in innate immune response, the activation effects of GDB on macrophages suggest that GDB may be a useful immunopotentiating agent.

A Clinical Study of Intercostal Neuropathy after Rib Fracture / 대한흉부외과학회지

BACKGROUND: The purpose of this study is to evaluate intercostal neuropathy after rib fracture and to determine the severity of intercostal neuropathy with using a numerical rating scale and according to the duration of pain and the body mass index. MATERIAL AND METHOD: We measured the positive sharp wave and fibrillation on the intercostal and paraspinal muscles in the thoracic region by performing needle electromyography in 47 patients who had intercostal neuralgia after rib fracture and who had needed daily analgesic for more than three months. RESULT: We diagnosed 11 cases as intercostal neuropathy among the 47 cases. Of the total 11 cases, 8 were male and 3 were female and they were most often of an active generation in the community. The common location of intercostal neuropathy was the intercostal space below the rib fracture and from the 7th to the 12th intercostal rib area. The incidence of intercostal neuropathy was significantly related with multiple rib fracture rather than single rib fracture. The symptoms observed were chest pain (90.9%), sensory change (81.8%), paresthesia and numbness (63.6%), back pain (27.2%) and muscle atrophy (18.2%). The numerical rating scale, the duration of pain and the body mass index showed no significant correlation with the severity of intercostal neuropathy. CONCLUSION: We concluded that the electrodiagnostic approach with considering the affecting factors and the clinical findings will be helpful for diagnosing and treating persistent intercostal neuralgic pain (more than 3 months) after rib fracture.

A Case of Primary Spontaneous Pneumothorax with a Three Nucleotide Deletion Mutation of the FLCN Gene / 대한흉부외과학회지

The cause of primary spontaneous pneumothorax (PSP) is obvious. Recently, the FLCN mutation was suggested to be a causal factor in PSP. A 47-year-old Korean male patient with chief complaint of repetitive PSP had numerous emphysematous bullae and multiple large cysts based upon high resolution computer tomography. Here we report a case of PSP with an FLCN c.468_470delTTC mutation.

Risk Factors for Recurrent Pneumothorax after Primary Spontaneous Pneumothorax / 대한흉부외과학회지

BACKGROUND: The purpose of this study was to identify factors associated with recurrent pneumothorax after wedge resection in primary spontaneous pneumothorax in our hospital. MATERIAL AND METHOD: Two hundred thirty-five consecutive patient (98% males; mean age, 23.9+/-4.5 years) who had undergone video-assisted thoracoscopic surgery (VATS) were reviewed retrospectively. The two groups were divided as follows: group A, non-recurrent patients (225 patients [96%]); and group B, recurrent group (10 patients [4%]); the risk factors were compared between the two groups. The single and multiple factors that influenced the recurrence rate were analyzed using Cox's proportional hazard model. RESULT: There were no significant differences between the recurrent and non-recurrent groups in terms of gender, smoking, site of recurrence, degree of collapse, operative time, and number or weight of resected bullae. The recurrence rate was significantly more common in the following: younger ages, increased height/weight ratio, longer initial air leakage period, and shorter duration of chest drainage. Early aggressive exercise (<30 days) of patients after wedge resection increased the tendency for recurrence. CONCLUSION: Thoracoscopic wedge resection does not have a higher recurrence rate than open thoracotomy. However, young age, height/weight ratio, continuous air, and duration of chest tube placement were risk factors for a recurrent pneumothorax.

Activation of Macrophage by Polysaccharide Isolated from Salicornia herbacea L / 체질인류학회지

Salicornia herbacea L. is one of the halophytes that can grow in salt marshes, or salt fields along the seashores in Korea. The objective of this study is to investigate the mechanism by which Salicornia Polysaccharide, (SPS) activates macrophages. To analyze macrophage activation and iNOS gene expression, we performed nitrite generation assay, immunohistochemistry, and reverse transcriptase-polymerase chain reaction. A polysaccharide isolated from the Salicornia herbacea L. significantly induces nitric oxide (NO). Immunohistochemical staining of inducible NO synthase (iNOS) showed that the increase of NO was due to the induction of iNOS production. RT-PCR analysis showed that SPS produced significant induction of iNOS gene expression. Immunohistochemical staining of p65 showed that SPS produced strong induction of NF-kappa B/Rel nuclear translocation. Electrophoretic mobility shift assay further confirmed the activation of NF-kappaB/Rel by SPS. In conclusion, we demonstrate that SPS stimulates the macrophages to express iNOS gene via the activation of NF-kappa B/Rel.

The Role of CD24 in Colonic Adenocarcinoma

PURPOSE: CD24 is a small, heavily glycosylated glycosylphosphatidylinositol-linked cell surface protein that is expressed in hematologic malignancies and in a large variety of solid tumors. It appears to function as a ligand of P-selectin, an adhesion molecule that is present in activated platelets and endothelial cells. We aimed to evaluate CD24 protein expression in adenomas and adenocarcinomas of the colon and to correlate it to clinicopathological data. METHODS: Adenomas and adenocarcinomas of the colon were stained for CD24 immunohistochemically. For statistical analysis, the staining was categorized according to stainability (negative, weakly, moderately, strongly positive) and staining patterns (membranous vs. intracytoplasmic). RESULTS: The present study clearly demonstrated that CD24 was much more abundantly expressed for adenocarcinomas than for adenomas in the colon (P <0.05). A higher significant association of cytoplasmic CD24 expression was observed with adenocarcinomas of the colon than with adenomas of the colon (P <0.05) and with positive nodal status of the colonic adenocarcinoma than with negative nodal status of the colonic adenocarcinoma (P <0.001). CONCLUSIONS: The stainability and the staining pattern of CD24 is an important molecular marker for colonic epithelial neoplasms and may help to define malignant transformation and to predict lymph-node metastasis.

Video-assisted Thoracoscopic Surgery in Posttraumatic Localized Clotted Hemothorax / 대한흉부외과학회지

BACKGROUND: Inadequate drainage of traumatic hemothoraces may result in prolonged hospitalization and complication such as empyema, fibrothorax and pleural calcification. This needs to be the placement of a tube thorascostomy which is efficacious in more than 80% of cases. Other cases require surgical treatment. MATERIAL AND METHOD: From March 2002 to February 2003, there were 123 patients who was done closed thorascostomy in traumatic hemothorax. 10 patients (group I) were undergone early retained clot evacuation with video assisted thoracoscopic surgery, but 5 patients (group II) who developed a localized hematoma or empyema were operated. Male were more than female and mean average was similar in both group. The most common cause of injury was traffic accidents and frequently combined lesions were a abdomen. RESULT: Interval from injury and operation, mean operation time, duration of tube drainage and hospital stay in group I were shorter than group II (p<0.05). Operation-related complication and recurrence of fluid collection within follow up period (17.8+/-3.8 months) in group I were none, but in group II (21.5+/-5.3 months) were 2 cases. CONCLUSION: Video assisted thoracoscopic surgery can be utilized as an effective and safe method for the removal of retained clotted hemothorax within 7 days.

Viscum Album Therapy in Malignant Pleural Effusion / 대한흉부외과학회지

BACKGROUND: Malignant pleural effusion is a common clinical problem in neoplastic patients. With the diagnosis of a malignant pleural effusion, palliative therapy was done. One of the treatments was a chemical pleurodesis. Talc was the most commonly used a sclerosing agent, but the quality of patient's life was not improved. We was evaluated by other agents such as Viscum album for relief of malignant pleural effusion. MATERIAL AND METHOD: From November 2001 to October 2003, 17 patients who underwent to chemical pleurodesis for the malignant pleural effusion. We compared the talc (group I: 10 patients) and Viscum album (group II: 7 patients). We analysed them retrospectively in term of various factors and results. RESULT: There were no significant differences between group I and group II in the sex ratio, mean age, origin of primary cancer and site, but, group I had higher successful rate (80%:71%) than group II. Group II had better length of chest tube stay after procedure, Karnofsky performance and recurrence than group I. The failed treatement group was related to the pleural fluid pH and interval of initial chemical pleurodesis after thoracostomy. CONCLUSION: Although the chemical pleurodesis with Viscum album was slightly lower than talc in the successful rate, there was an alternative method instead of the chemical pleurodesis with talc to improve the patient's quality of life in malignant pleural effusion.

Video-assisted Thoracoscopic Surgery in Posttraumatic Localized Clotted Hemothorax / 대한흉부외과학회지

BACKGROUND: Inadequate drainage of traumatic hemothoraces may result in prolonged hospitalization and complication such as empyema, fibrothorax and pleural calcification. This needs to be the placement of a tube thorascostomy which is efficacious in more than 80% of cases. Other cases require surgical treatment. MATERIAL AND METHOD: From March 2002 to February 2003, there were 123 patients who was done closed thorascostomy in traumatic hemothorax. 10 patients (group I) were undergone early retained clot evacuation with video assisted thoracoscopic surgery, but 5 patients (group II) who developed a localized hematoma or empyema were operated. Male were more than female and mean average was similar in both group. The most common cause of injury was traffic accidents and frequently combined lesions were a abdomen. RESULT: Interval from injury and operation, mean operation time, duration of tube drainage and hospital stay in group I were shorter than group II (p<0.05). Operation-related complication and recurrence of fluid collection within follow up period (17.8+/-3.8 months) in group I were none, but in group II (21.5+/-5.3 months) were 2 cases. CONCLUSION: Video assisted thoracoscopic surgery can be utilized as an effective and safe method for the removal of retained clotted hemothorax within 7 days.

Viscum Album Therapy in Malignant Pleural Effusion / 대한흉부외과학회지

BACKGROUND: Malignant pleural effusion is a common clinical problem in neoplastic patients. With the diagnosis of a malignant pleural effusion, palliative therapy was done. One of the treatments was a chemical pleurodesis. Talc was the most commonly used a sclerosing agent, but the quality of patient's life was not improved. We was evaluated by other agents such as Viscum album for relief of malignant pleural effusion. MATERIAL AND METHOD: From November 2001 to October 2003, 17 patients who underwent to chemical pleurodesis for the malignant pleural effusion. We compared the talc (group I: 10 patients) and Viscum album (group II: 7 patients). We analysed them retrospectively in term of various factors and results. RESULT: There were no significant differences between group I and group II in the sex ratio, mean age, origin of primary cancer and site, but, group I had higher successful rate (80%:71%) than group II. Group II had better length of chest tube stay after procedure, Karnofsky performance and recurrence than group I. The failed treatement group was related to the pleural fluid pH and interval of initial chemical pleurodesis after thoracostomy. CONCLUSION: Although the chemical pleurodesis with Viscum album was slightly lower than talc in the successful rate, there was an alternative method instead of the chemical pleurodesis with talc to improve the patient's quality of life in malignant pleural effusion.

Clinical Analysis of Mitral Valve Repair with Artificial Chordae / 대한흉부외과학회지

Background: Failure of mitral valve repair sometimes may be ascribed to severe or progressive alteration of the subvalvar apparatus. The aim of this study was to evaluate the effects of new chordae formation on mitral repair. Material and Method: From March 1997 to February 1999, 26 patients underwent mitral valve repairs with new chordae formation, we compared the symptoms and echocardiographic findings checked at preoperative state, and intraoperative period, discharge, and their last OPD visit. There were 15 male , and 11 female patients, and their mean age was 51.2+/-13.4 years. Etiology of the lesions was degenerative (18), rheumatic (6), infective (1) and ischemic (1). Chordal lesions were caused by rupture (18), elongation (6), and a combination of two causes (2). Associated lesions included atrial septal defect (2), tricuspid insufficiency (7), aortic insufficiency(4), and a combination of previous two factors (2). The number of mean artificial chordae was 3.6+/-1.6. Annuloplasty was performed in all cases. The CPB time was 182.1+/-63.7 minutes and the ACC time was 133.1+/-45.6 minutes. Average follow up period was 49.2+/-7.1 months. Result: There was no early death. Early reoperation was performed in two patients, one patient received mitral valve replacement because of an abnormality of annuloplasty and another received pericardiostomy due to postoperative pericardial effusion. During the follow up of 49.2+/-7.1 moths, there was no late mortality. Postoperative NYHA functional class checked at last OPD visit was class I in 22 patients (88%), class II in 2 (8%), and class III in 1 (4%). Regarding the late echocardiogram MR was absent in 20 patients (78%), I in 4 (15%), and II in 1 (4%). The postrepair mitral valve area was 2.2+/-0.35 cm(2). Conclusion: This study suggests that mitral valve repair using new chordae formation provides good early and mid term survivals and functional improvement. We think that the artificial chorda formation with polytetrafluoroethylene suture might be safe and effective technique for mitral valve repair.

Isolated Right Ventricular Hypoplasia: A case report / 대한흉부외과학회지

Isolated right ventricular hypoplasia is a rare clinical entity. We describe a case of right ventricular hypoplasia, single atrium and spongy myocardium of left ventricle. The volume of right ventricle was half the volume of left ventricle and z-value of tricuspid valve was -4 preoperatively. The patient, 6-year-old boy, underwent atrial partitioning with 3 mm fenestration. Postoperative course was smooth and he tolerated the biventricular state well during follow-up. Follow-up catherterization was done 27 months later. The tricuspid valve grew well (z-value=-0.4) and atrial septal fenestration is closed spontaneously. This article reports a case of successful biventricular repair in a patient with isolated right ventricular hypoplasia.

A Experimental Study of PTEN (Phosphatase and Tensin) Role in Mesothelioma / 대한흉부외과학회지

BACKGROUND: Conventional treatment for mesothelioma is largely ineffective. We evaluated the novel approach of adenoviral gene transfection of PTEN gene in mesothelioma cancer cell lines, inflammatory and epithelial subtype, which are sensitive to adenoviral p53. MATERIAL AND METHOD: Binary adenoviral PTEN and LacZ (Ad/GT-LacZ and Ad/GV16) vectors were used for transduction of the mesothelioma cell lines, REN (p53 sensitive). Protein levels were determined by Western blotting assay. Apoptosis was assessed by fluorescence-activated cell sorter analysis of subdiploid populations. Cell viability was determined with the XTT assay. Statistical analysis was performed with analysis of variance and the Student t test. RESULT: 72 hours after the treatment of adenoviral PTEN gene, cell killing were 32.9% for REN compared to control cell (2.5%) at MOI of 20. Also we observed the over-expression of proapoptotic protein, bax and decreased expression of bcl-2 protein in REN cells. But the expression of BCL-xl, Bak, Bad proteins were not altered. CONCLUSION: Adenovirus Pten-mediated overexpression of the Bax gene induces apoptosis and decreased cellular viability in p53-sensitive mesothelioma cells. These data suggest that the transfection of PTEN gene may represent a alternative gene therapy strategy to treat mesothelioma.

Norwood Procedure with Home-made Bovine Pericardial Patch: A report of 2 cases / 대한흉부외과학회지

A patch design using bovine pericardial patch for aortic enlargement in the Norwood procedure has been introduced to avoid problems related to homograft availability. We report 2 successful cases of Norwood procedure with home-made bovine pericardial patch. The first case was a 23-day-old (2.2 kg) patient with multi-level left ventricular outflow tract obstruction with ductal-dependent systemic circulation. The other case was a 9-day-old (3 kg) patient with hypoplastic left heart syndrome. This technique was relatively easy to perform, reproducible and quite effective like homograft patch.

A New Technique of Angioplasty of the Left Pulmonary Artery Using an Autologous Main Pulmonary Artery Flap / 대한흉부외과학회지

BACKGROUND: Stenosis of the left pulmonary artery (LPA) after repair of tetralogy of Fallot (TOF) is troublesome. A new technique of LPA angioplasty using an autologous MPA flap was performed in patients with TOF. MATERIALAND METHOD: From October 1998 to January 2001, 24 patients (median age; 10 months, range; 4 to 145 months)underwent total correction of TOF with LPA angioplasty using the autologous MPA flap. Five patients underwent pulmonary angioplasty without any patch over the MPA and LPA. The patches were required to enlarge only the MPA in 4 patients, and transannular RVOT widening was performed in 15. RESULT: There were no operative or late deaths. During follow-up (range: 6~42 months), reoperation for LPA stenosis was not required in any patients, but balloon angioplasty for branch pulmonary artery stenosis was performed in 3 patients. Echocardiography and CT angiography at the recent follow-up showed an obtuse angle between the MPA and LPA. CONCLUSION: Although further follow-up is needed, the angioplasty using the autologous MPA flap can be easily performed, avoiding patch-related complications, and allowing growth of the MPA flap. This angioplasty technique creates a more natural and obtuse angle between the MPA and LPA, which can minimize kinking of the LPA, especially in the patients who underwent transannular patch widening.

Biventricular Repair of Critically Ill Neonate with Ebstein's Anomaly: Report of 1 case / 대한흉부외과학회지

Patients with severe Ebstein's anomaly showing in the neonatal period, represent progressive cardiac enlargement with pulmonary hypoplasia and functional pulmonary atresia with patent ductus arteriosus-dependent pulmonary circulation. Biventricular repair in these patients had been mostly unsuccessful except for Starnes' procedure that converts the anatomy to single ventricle physiology for Fontan procedure. A 4-days old male was admitted with the diagnosis of severe Ebstein's anomaly with anatomic pulmonary atresia and severe cardiac enlargement. He successfully underwent biventricular repair with vertical plication method of atrialized right ventricle, tricupid annuloplasty, transannular right ventricular outflow tract reconstruction, atrial septal defect patch closure with fenestration, and right atrial reduction angioplasty. Postoperatively, cardiothoracic ratio was significantly reduced and mild tricuspid regurgitation was remnant in echocardiography. The patient is currently 10 months old and is fully active without restrictions.