The Her-2/neu protooncogene encodes a transmembrane tyrosine kinase that is structurally homologous to the receptor for epidermal growth factor. Its amplification and overexpression are associated with poor prognosis in breast cancer patients. Neu differentiation factor is a ligand for Her-2/neu protooncogene and was detected in ras-transformed rat fibroblasts. Heregulin (human homologue of neu differentiation factor) is a 44-kilodalton glycoprotein that stimulates tyrosine phosphorylation and induces growth arrest or stimulation and differentiation in human breast cancer cell lines. In this study we examined the expression of heregulin mRNA by nested reverse transcription (RT) PCR with fresh tissue, Her-2/neu protein, ICAM-1 and steroid receptors by immunohistochemistry, and DNA ploidy pattern by flow cytometry with paraffin-embedded tissue in invasive breast carcinoma. We compared the data with nodal status, lymphovascular invasion, steroid receptor status and DNA ploidy pattern. For RT-PCR to heregulin mRNA, 38 cases of fresh breast cancer tissue were obtained. Total 68 cases of invasive breast carcinoma tissue were fixed in formalin, which were used for routine histology, immunohistochemistry and flow cytometry. The results are as follows; 1) Heregulin mRNA was expressed in 86.1% of patients with invasive breast carcinoma and 100% of patients with benign breast lesion using nested RT-PCR analysis. 2) Her-2/neu protein was overexpressed in 50.0% of tumors using immunohistochemistry. The expression of Her-2/neu protein was significantly correlated with high counts of lymph nodes with metastasis (p<0.05), and high nuclear grade (p<0.05). 3) Her-2/neu protein overexpression was significantly correlated with a high DNA index(p<0.05). All of the tumors showing Her-2/neu protein overexpression and no heregulin mRNA expression revealed near tetraploid DNA content. However, both Her-2/neu overexpression and heregulin mRNA expressing tumors revealed near tetraploidy in 38.9% and diploidy in 50.0%. Based on these results, heregulin mRNA expression rate was 86.1% in human invasive breast carcinoma. Her-2/neu protein overexpression is associated with high positive lymph node number and DNA index. Statistically significant reverse correlation with lymph node metastasis is not present.
Animals ; Breast Neoplasms* ; Breast* ; Cell Line ; Diploidy ; DNA* ; Epidermal Growth Factor ; Fibroblasts ; Flow Cytometry ; Formaldehyde ; Glycoproteins ; Humans* ; Immunohistochemistry ; Intercellular Adhesion Molecule-1 ; Lymph Nodes ; Neoplasm Metastasis* ; Neuregulin-1* ; Phosphorylation ; Ploidies* ; Polymerase Chain Reaction ; Prognosis ; Protein-Tyrosine Kinases ; Rats ; Receptors, Steroid ; Reverse Transcription ; RNA, Messenger* ; Tetraploidy ; Tyrosine

PURPOSE: The CD5 molecules are pan-T cell antigens and are found on a minor subpopulation of B cells. CD5 antigens are involed in an intracellular signal transduction as well as in an intercellular signal transduction between CDS+ T cell/CD72+ B cell by CD5/CD72 interaction. CD5 antigens are known to be participated in classic immune reactions and in this study CDS mRNA expressions by lymphocytes were examined in allergic patients controls, acute febrile infectious disease controls and normal controls to elucidate the possibility of CDS involvement in allergic immune reactions. METHODS: Fifteen allergic patients, ten patients of acute febrile infectious disease patients and ten normal controls were studied. Venous blood was drawn and mononuclear cells were separated. T cells and B cells were separated using immunomagnetic beads. Total RNA was extracted and RT-PCR (reverse transcriptase - polymerase chain reaction) was done to detect CDS antigen mRNA expression. RESULTS: 1) CDS mRNA overexpressions were detected in allergic patient controls as compared to that in acute febrile infectious controls. CDS mRNA was not detected in normal controls. Semiquantitative CD5 mRNA expressions were measured as relative expressions of CD5 to GAPDH. Relative quantities of CD5 mRNA expressions were 90.656.24% in allergic patient controls and 23.76+3.58% in acute febrile infectious patients. CONCLUSIONS: CDS mRNA overexpression is a characteristic phenomenon in allergic immune reactions. From these result, CD5/CD72 pathway might be the preference immune mechanism in allergic immune reaction and the further study for the exact mechanism of CDS involvement in allergic immune reactions may be necessary
Antigens, CD5 ; B-Lymphocytes ; Communicable Diseases ; DNA-Directed RNA Polymerases ; Humans ; Hypersensitivity ; Lymphocytes* ; RNA ; RNA, Messenger* ; Signal Transduction ; T-Lymphocytes

We examined C3H pregnant mice at 15 days (70% gestation) after treatment of lipopolysaccaride (LPS) to observe the changes of IL-6 concentration in maternal serum and amniotic fluid and expression of IL-6, IL-13 & TIMP-3 in placenta, fetus and endometrium, and to investigate the correlation among IL-6, IL-13 and TIMP-3. The results were as follows: 1) IL-6 in serum and amniotic fluid after treatment of LPS was significantly elevated; peaked at 1, 2, 4, 5 hours and decreased to control level at 8 hours (P<0.05). IL-6 in placental disc, chorioamnionic membrane, fetus, decidua and endometrial epithelium was overexpressed significantly at 1, 2, 4 hours after treatment of LPS (P<0.05). IL-6 overexpression was more significantly increased in maternal tissue than fetal tissue (P<0.05). 2) Increased concentration of amniotic fluid IL-6 was equally originated from transplacental crossage of maternal serum IL-6, and direct local production of IL-6 from placenta, fetus and endometrium (P<0.05). 3) IL-13 in placental disc, chorioamnionic membrane, fetus, decidua and endometrial epithelium was overexpressed after treatment of LPS, but not significant statistically. 4) TIMP-3 was overexpressed in placental disc, chorioamnionic membrane, fetus and decidua. TIMP-3 overexpression was more significant in placental disc than other tissues (P<0.05). 5) Overexpressions in IL-13 and IL-6 revealed direct proportional correlation coefficient (Spearman correlation coefficient, 0.5212 ; P<0.05). IL-6 expression was a head of overexpression of TIMP-3, but not significant. In conclusion, all of IL-6, IL-13 and TIMP-3 relate with inflammatory response, especially IL-6 in maternal serum, amniotic fluid and tissue of placenta, fetus and endometrium was so sensitive that it can be an indicator for antenatal diagnosis of chorioamnonitis, and amniotic fluid IL-6 is equally originated from maternal serum and from tissue of placenta, fetus and endometrium. IL-13 and TIMP-3 may have parallel correlation to the IL-6 in fetal and maternal tissue after treatment of LPS.
Amniotic Fluid ; Animals ; Decidua ; Endometrium* ; Epithelium ; Female ; Fetus* ; Head ; Interleukin-13* ; Interleukin-6* ; Membranes ; Mice* ; Placenta* ; Prenatal Diagnosis ; Tissue Inhibitor of Metalloproteinase-3*

Telomerase is an enzyme that maintains telomeres and prevents telomere shortening, and may be linked with cellular proliferation or the aging process. The purpose was to examine telomerase activity in human chorionic villi from early and term normal pregnancies, and to analyze the correlation of telomerase activity (TA) with MIB-1 & bcl-2. A total of 37 placentae were obtained from 16 early and 21 term pregnancies. TA was assayed by telomeric repeat amplification protocol, and immunohistochemical staining was performed for MIB-1 & bcl-2 expression. TA & MIB-1 expression were strong in early placenta, but bcl-2 was highly expressed in term placentae. Thirteen (81.25%) of 16 early placentae showed TA, but only 2 (9.52%) of 21 term placentae expressed TA (p<0.01). MIB-1 was observed in nuclei of cytotrophoblast, and the expression rate was 16.09% in early placentae and 2.87% in term placentae (p<0.01). bcl-2 was observed only in the cytoplasm of syncytiotrophoblast. Term placenta demonstrated stronger expression of bcl-2 compared to early placentae (p<0.05). These findings suggest that TA, MIB-1 & bcl-2 expression are critically regulated over the course of gestation: cytotrophoblast, main cells of early chorionic villi, may be a common source of telomerase and proliferative activity. The TA showed good correlation with cellular proliferative activity. Syncytiotrophoblast, may be a main source of bcl-2 expression which is stronger in the term placentae.
Aging ; Cell Proliferation ; Chorion* ; Chorionic Villi* ; Cytoplasm ; Humans* ; Placenta ; Pregnancy* ; Telomerase* ; Telomere ; Telomere Shortening ; Trophoblasts

Spinal Anesthesia employing 0.5% plain bupivacaine was administered to 40 patients scheduled for lower limb or perineum sThe plasma concentrations and pharmacokinetic parameters of lidocaine were studied in 4 patients under general anesthesia(halothane, or enflurane-N20-O2) following the introduction of an 1% lidocaine endotracheal spray(1.5mg/kg) through an epidural catheter. Poak plasma lidocaine levels were reached in 5 to 15 minutes and were within the nontoxic range. The pharmacokinetics of lidocaine in these patients can be described by a two-compartment model with a rapid alpha distribution(T 1/2 alpha 8.66+/-2.24 min.), and an extensive apparent volume of idstribution(1.32+/-0.46 1/kg) similar to that observed in normal subjects. The half-life of absorption was 3.65+/-1.21 minutes. However, the elimination half-life(T 1/2 beta 173.25+/-32.41 min.) was prolonged and the total plasma clearance( 6.15+/-3.25 ml/min/kg) was decreased. This potent inhalation anesthetic agent may reduce the hepatic blood flow and would be expected to reduce the plasma clearance of lidocaine by reducing the delivery of plasma lidocaine. This study suggests that tracheal administration of lidocaine will produce effective plasma lidocaine levels in many clinical situations as will intravenous administration.
Absorption ; Administration, Intravenous ; Anesthesia, General* ; Anesthesia, Spinal ; Bupivacaine ; Catheters ; Half-Life ; Humans ; Inhalation ; Lidocaine* ; Lower Extremity ; Perineum ; Pharmacokinetics* ; Plasma

Warfarin is a frequently prescribed anticoagulant in rehabilitation patients. Adverse drug reactions of warfarin were reported as bleeding and cutaneous microvascular thrombosis. Major bleeding, such as intracranial hemorrhage and psoas hematoma, in patients receiving anticoagulation therapy is a rare condition, but sometimes very serious complication that can even be fatal. Patient-specific factors (eg, age, body size, race, concurrent diseases, and medications) explain some of the individual variability in warfarin dose, but genetic factors, which influence warfarin response, explain a significantly higher proportion of the variability in the dose. There are two identified genes that are responsible for the main proportion of the genetic effect: CYP2C9, which codes for the enzyme cytochrome P450 2C9 that metabolizes S-warfarin, and VKORC1, which codes for warfarin's target, vitamin K epoxide reductase. We report a case of intolerance to warfarin dosing, due to impaired drug metabolism in a patient with CYP2C9*1/*3 and VKORC 1173TT. Fortunately, there are no severe complications.
Body Size ; Continental Population Groups ; Cytochrome P-450 Enzyme System ; Drug Toxicity ; Hematoma ; Hemorrhage ; Humans ; Intracranial Hemorrhages ; Mixed Function Oxygenases ; Oxidoreductases ; Thrombosis ; Vitamin K ; Warfarin

No abstract available.
Polymerase Chain Reaction*

The acquisition of human chondrocytes for transplantation and cartilage coverage presents a major problem as these cells dedifferentiate rapidly during expansion in monolayer culture. Dedifferentiated chondrocytes change their shapes, metabolic states, and programs of matrix biosynthesis. We initiated this study on the basis of the hypothesis that bFGF, VEGF, and micromass culture can influence both the proliferation and their ability to express COL2A1 gene as a chondrogenic marker and Cbfa1 gene as an osteogenic marker. Chondrocytes in monolayer and micromass culture with or without bFGF and VEGF in vitro were collected and analyzed. In results, bFGF stimulated the proliferation of chondrocytes in monolayer culture. VEGF also stimulated the proliferation, but was less effective. The phenotype of chondrocytes was gradually changed in monolayer culture. Chondrocytes expanded in the presence of bFGF became dedifferentiated. However, dedifferentiated chondrocytes fully maintained their potential for redifferentiation in response to environmental changes. After transferring in micromass culture, chondrocytes which expanded with bFGF demonstrated high COL2A1 expression that was biochemically comparable to primary chondrocytes. Chondrocytes which expanded with VEGF demonstrated high Cbfa1 expression in both monolayer and micromass culture with passage times. This study provides that bFGF is needed to expand chondrocytes during tissue cultivation and additional three-dimensional environment is needed to maintain their differentiated phenotype. VEGF initiates the osteogenic potential during the chondrocyte expansion especially in micromass culture.
Cartilage ; Chondrocytes* ; Humans* ; Phenotype ; Vascular Endothelial Growth Factor A*

This study investigated the expression of osteopontin (OPN) in rat lumbar spinal cords after lumbar nerve root avulsion, using in situ hybridization histochemistry, immunocytochemistry and western blot analysis. Cells expressing OPN were motoneurons and interneurons in the ventral horn, but no signals were observed in neurons in the dorsal horn of the normal lumbar spinal cord. From day 1 after avulsion injury, OPN mRNA-labeled neurons increased in the ventral horn and the intermediate zone. By day 3, relatively strong OPN mRNA signals were found throughout the gray matter of the injured side of the spinal cord with OPN mRNA-labeled cells scattered in the superficial dorsal horn. By day 7, the labeling patterns for OPN mRNA were similar to those on day 3, but the numbers of OPN mRNA-labeled cells in the ventral horn and the intermediate zone peaked. At this point, these labeled cells were also more densely packed and the intensity of signals was stronger. Interestingly, these labeled cells were neurons, but not glial cells such as astrocytes or microglia. This OPN mRNA-labeled cell profile in the dorsal horn had nearly disappeared by day 14 after avulsion injury, and the labeling pattern became similar to that on day 1. By day 28, after avulsion injury, the numbers of OPN mRNA-labeled cells decreased further below control values. These results suggest that increased expression of OPN in the rat lumbar spinal cord after avulsion injury might play an important role in the pathogenesis of damaged neurons.
Animals ; Astrocytes ; Blotting, Western ; Horns ; Immunohistochemistry ; In Situ Hybridization ; Interneurons ; Microglia ; Neuroglia ; Neurons ; Osteopontin* ; Radiculopathy ; Rats* ; RNA, Messenger ; Spinal Cord*

Acetylsalicylic acid (ASA) is one of the most commonly used medications in global market, with a risk of intoxication in certain patients. However, monitoring blood drug concentration often requires frequent hospital visits; hence there is an unmet need to increase patientcentricity by conducting blood sampling at home. Volumetric absorptive microsampling (VAMS) is a device that allows collection of homogenous and accurate volume of blood without venipuncture, and can be utilized by patients who are not in hospital settings; but because ASA is prone to hydrolysis and stabilizing reagents cannot be added to VAMS samples, a way to improve sample stability must be developed. The objective of this study was to identify the cause of instability with ASA samples collected by VAMS, and to evaluate ways to improve sample stability. A liquid chromatography with tandem mass spectrometry (LC-MS/MS) was used for analysis of ASA concentration in whole blood. Samples collected with VAMS were kept under different drying conditions (desiccator, pressurized, nitrogen gas and household vacuum sealer) and were compared to the control samples collected by conventional venous sampling. The recovery of ASA was about 31% of the control when VAMS sample was dried at room temperature, whereas VAMS samples under humidity controlled conditions showed more than 85% of recovery. Our results suggest that adequate level of humidity control was critical to ensure sample stability of ASA, and this humidity control could also be achieved at home using household vacuum sealer, thus enabling patient-centric clinical trials to be conducted.