Cytokines and hematology.
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Bone Marrow Transplantation↗
Biomedical subjects
S Kasakura
Publications and source records attributed to S Kasakura.
Hypocoagulable state of human preovulatory ovarian follicular fluid: role of sulfated proteoglycan and tissue factor pathway inhibitor in the fluid.
Ovulation accompanied by tissue damage can cause an increase in the level of tissue factor (TF) in the follicular fluid, triggering the extrinsic coagulation pathway. However, follicular fluid must block fibrin formation and maintain fluidity until the release of the oocyte at ovulation. The combination of sulfated proteoglycan, antithrombin, and TF pathway inhibitor (TFPI) appears to play a critical role in the hypocoagulability of human follicular fluid. When compared with plasma, folicular fluid differs markedly in the levels of a number of important coagulation proteins. Principal among these are 15-fold, 13-fold, and 3.7-fold increases in free TFPI, thrombin-antithrombin complex, and TF, respectively. The excessively prolonged activated partial thromboplastin time (APTT) and prothrombin time (PT) of human ovarian follicular fluid appear to be primarily due to high concentrations of sulfated proteoglycans, which accelerate the inactivation of thrombin and the anti-Xa activity of TFPI. Thus, heparitinase treatment shortened the clotting times of follicular fluid and reduced the inhibition of thrombin by the proteoglycan fraction combined with a fraction containing antithrombin. The remaining prolongation of APTT and PT may be caused by high levels of free TFPI in follicular fluid, which were confirmed by Northern blotting analysis, demonstrating TFPI mRNA expression by granulosa cells.
Antithrombin III↗
Creatine kinase gene mutation in a patient with muscle creatine kinase deficiency.
BACKGROUND: We describe a 56-year-old woman admitted to the hospital with a diagnosis of acute myocardial infarction without an increase of serum creatine kinase (CK) activity during her clinical course. She died on the 11th hospital day, and the diagnosis was confirmed by autopsy. The patient had had no previous muscular symptoms. METHODS: Expression of the CK-muscle (CK-M) protein in cardiac tissue was examined by immunoblotting and immunochemical staining. CK-M mRNA expression was estimated by semiquantitative reverse transcription-PCR. Gene structure of CK-M was determined by Southern blotting and direct sequencing of 2251 bp. Existence of a point mutation in the CK-M gene was examined by restriction fragment length polymorphism analysis of PCR products (PCR-RFLP) in the patient and in 108 controls. RESULTS: CK-M protein in the myocardial tissue of the patient was substantially lower (103 +/- 7 ng/mg protein) than in control myocardial tissue (35 800 +/- 2860 ng/mg protein). Immunoreactive CK-M in the patient tissue sample was 0.3% of the value for the control sample. CK-M mRNA was 53-fold less in the patient sample compared with the control. This very low expression of CK-M mRNA was considered to be the primary reason for CK-M deficiency. Direct sequencing revealed a point mutation at residue 54 in exon 2, which was specific for the patient. No other abnormalities were found in the CK-M gene of the patient. CONCLUSIONS: This report identifies a molecular abnormality in human CK deficiency and discusses the physiologic relevance of CK-M.
Blotting, Southern↗
Induction of follicular gastritis following postthymectomy autoimmune gastritis in Helicobacter pylori-infected BALB/c mice.
Helicobacter pylori is the major causative agent of chronic antral gastritis and is thought to be involved in the pathogenesis of mucosa-associated lymphoid tissue lymphoma (MALToma) developing in the human stomach. The aim of this study was to clarify whether corporal autoimmune gastritis (AIG), which is known to decrease acidity due to destruction of parietal cells, predisposes mice to H. pylori infection, thereby leading to MALToma-like pathology. BALB/c mice in which AIG had been induced by thymectomy 3 days after birth (AIG mice) were used. The AIG mice were orally administered mouse-adapted H. pylori at the age of 6 weeks and were examined histologically and serologically after 2 to 12 months. The results were compared with those obtained from uninfected AIG mice and infected normal mice. Germinal centers were induced in the corpus in 57% of the H. pylori-infected AIG mice, which elicited anti-H. pylori antibody responses in association with upregulation of interleukin-4 (IL-4) mRNA. In these mice, parietal cells remained in the corpus mucosa. These findings were in contrast to those with the uninfected AIG mice: fundic gland atrophy due to disappearance of parietal cells associated with upregulation of gamma interferon, but not IL-4, mRNA and no germinal center formation in the corpus. These observations suggest that AIG alters the infectivity of H. pylori, leading to MALToma-like follicular gastritis, at an early stage after H. pylori infection.
Animals↗
[Extraordinary high elevation of serum CA19-9 levels in an apparently healthy subject].
We studied the woman presenting extraordinarily high serum CA19-9 levels who did not have any disease that was known to result in elevation of the serum CA19-9 level. Her CA19-9 values in sera obtained by 16 determinations with the enzyme immunoassay (EIA) over a period of 7 years were from 820 U/ml to 1,310 U/ml (median 1,015 U/ml). The correlation was good between serum CA19-9 levels in this subject with the EIA and radio immunoassay (RIA). The molecular weight of CA19-9 was determined by high-performance liquid chromatography (TSK gel G4000 SWXL column). While the molecular weights of CA19-9 in each patient with pancreatic cancer, cholangioma, cholecystitis or pancreatitis that were used as controls, were greater than 1,000 kDa, the CA19-9 of this subject had the molecular weight of approximately 550 kDa. To further determine whether human immunoglobulin was involved in the elevation of CA19-9 in the serum of this subject, we investigated the effect of adding rabbit anti-human IgM, IgG and IgA sera to the serum of this subject, CA19-9 values were unaffected by these antisera. Thus, the data obtained from the absorption study indicated that an anti-idiotypic antibody against the anti-CA19-9 antibody is unlikely to be responsible for an elevation of CA19-9 in the serum of this subject.
Antibodies, Anti-Idiotypic↗
[Cytokines and their receptors].
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Cytokines↗
[Clinical usefulness of lectin-reactive fraction of alpha-fetoprotein in hepatocellular carcinoma].
Alpha-fetoprotein (AFP) is well known as a tumor marker of hepatocellular carcinoma (HCC). AFP shows micro heterogeneity because of a structural variance in its sugar chain, and the sugar chain obtained from HCC patients has a high affinity to lectins such as Lens culinaris agglutinin. The lens culinaris agglutinin A-reactive fraction of alpha-fetoprotein (AFP-L3) was detected by a sensitive method using lectin-affinity electrophoresis coupled with antibody-affinity blotting. Of 62 HCC patients examined before initial therapy, 23 patients (37.1%) had an AFP-L3 level greater than 15%. In contrast, none of the patients with liver cirrhosis or chronic hepatitis tested were positive for AFP-L3. Seven (26.9%) of 26 patients with small HCC less than 2 cm in diameter measured on ultrasonography or computed tomography showed elevated AFP-L3 levels. Because there was no correlation observed between AFP-L3 and PIVKA-II, the combined measurement of these two complementary markers appeared to be useful in the diagnosis of HCC. When the results of both assays were combined, 34 (54.8%) of the 62 patients with untreated HCC showed elevated levels of one or both markers. The use of both markers together with imaging modalities during regular follow-up of chronic liver disease patients who are at high risk for HCC might be of great benefit.
Antibody Affinity↗
[A role for T-helper type 1 and type 2 cytokines in the pathogenesis of various human diseases].
Mosmann first proposed the existence of subsets of CD4+ T cells that produce distinct types of cytokines. Native T lymphocytes (Thp cells) differentiates into either CD4+ Th1 cells that produce IL-2, IFN gamma, and lymphotoxin which promote cell-mediated immunity, or into Th2 cells that produce IL-4, IL-5, IL-6, IL-10 and IL-13, which promote antibody production and humoral immunity. These T cell subsets reciprocally regulate one another since one of the Th1 products, IFN gamma, inhibits the proliferation and functions of Th2 cells, whereas the Th2 products, IL-4 and IL-10, suppress cytokine production by Th1 cells. A distinct Th1/Th2 divergence determine resistance versus susceptibility to diseases such as leishmaniasis and toxoplasmosis in mice. In allergic diseases such as atopic dermatitis and allergic asthma, allergen-specific T cells acquired the Th2 phenotype. These Th2 cells produce IL-4, IL-5, IL-6, IL-10 and IL-13. These cytokines induce eosinophilia and an Ig class switch to IgG4 and IgE. These Th2 cells are responsible for the enhanced production of IgE antibodies. These findings indicate that Th2 cytokines play an important role in the development of allergic diseases. The importance of cell-mediated immunity, particularly donor-anti-host CTL, in mediating acute GVHD suggests that Th1 cytokines may be important in the induction of acute GVHD. To further characterize the roles of Th1 and Th2 cytokines in the development of acute GVHD, analysis of IL-2, IFN gamma, IL-4 and IL-10 cytokine genes was performed by RT-PCR on biopsied skin specimen. An increase in mRNA expression for IL-2 and IFN gamma was observed, whereas there was no significant increase in IL-4 and IL-10 mRNA. These data suggest that Th1 cytokines may be essential for the development of acute GVHD. It is apparent that Th1 cytokines are generally harmful to the maintenance of pregnancy. We have shown that Th2 cytokines are produced by maternal T lymphocytes at the maternal-fetal surface (retroplacental blood lymphocytes). This finding strengthens the hypothesis of a significant contribution of Th2 cytokines to a successful pregnancy.
Animals↗
[Clinical usefulness of serum PIVKA-II levels determined by ECLIA system as a tumor maker for hepatocellular carcinoma].
PIVKA-II is well known as a tumor maker of hepatocellular carcinoma (HCC). We measured serum PIVKA-II concentrations with a commercially available PIVKA-II immunoassay kit (Picolumi PIVKA-II: Eisai Co., Ltd., Tokyo) using Electrochemiluminescence Immunoassay (ECLIA). ECLIA system is a novel immunoassay system using a Ruthenium (II) Tris (bipyridyl) luminesced by electric energy. Cut off value was 40 mAU/ml by receiver-operating characteristic curves as a tumor maker for HCC. Eighty-nine out of 142 (62.2%) patients with HCC had elevated serum PIVKA-II levels and seventeen out of 36 (47.2%) patients whose tumor size was below 2 cm in diameter showed high PIVKA-II levels. We determined the serial changes in serum PIVKA-II levels of two patients with HCC following initial therapy. In these patients, elevations of serum PIVKA-II levels determined by ECLIA system preceded the HCC relapse detected by imaging diagnostic procedures. In summary, this assay system is suitable for detecting small increases in PIVKA-II concentrations. Determination of PIVKA-II by this assay system is found to be useful for the early detection of HCC.
Aged↗
Structure of the TCR expressed on a gastritogenic T cell clone, II-6, and frequent appearance of similar clonotypes in mice bearing autoimmune gastritis.
A parietal cell-specific Th1 clone, II-6, which was established from a BALB/c mouse bearing post-thymectomy autoimmune gastritis (AIG), recognizes a peptide of the alpha subunit (alpha891-905) of H+/K+-ATPase and induces gastritis in nu/nu BALB/c mice by adoptive cell transfer. In the present study, the primary structure of the TCR of II-6 was determined as Valpha10-Jalpha c5a-Calpha and Vbeta14-Jbeta2.3-Cbeta2 by cDNA cloning. Using PCR with specific primers, we defined the use of this II-6 TCR in nu/nu mice with transferred II-6 cells and in mice that spontaneously developed AIG by thymectomy on day 3 after birth (d3-Tx). II-6 TCR mRNAs were detected in the gastric mucosa of all of the nu/nu mice, suggesting that II-6 cells indeed home to the gastric mucosa and thereby were directly involved in the destruction of target parietal cells. TCR beta chain mRNAs encoding CDR3 region sequences almost identical with that of II-6 were also found in the gastric mucosa in 43% (six of 14 mice tested) of the d3-Tx AIG mice at 4-12 weeks old by nested RT-PCR. Such a frequent appearance of similar clonotypes in independent individuals suggests that T cells bearing II-6-like TCR including the II-6 itself might be directly involved in, although not essential for, the pathogenesis of AIG in 3d-Tx mice.
Amino Acid Sequence↗
[Study of serum thrombomodulin(TM) levels in patients with hyper- or hypo- thyroidism].
We studies a relationship between the serum levels of thrombomodulin(TM) and the thyroid functions. Serum TM levels were measured in 48 patients with Graves' disease, 17 patients with primary hypothyroidism, 7 patients with subacute thyroiditis, 5 patients with painless thyroiditis and 2 patients with systematic Refetoff syndrome. These patients did not have malignant tumor, kidney failure, or blood vessel injury. Control sera were obtained from 42 healthy subjects. Serum levels of TM in patients with untreated Graves' disease were significantly higher(p < 0.001) compared with those in controls. Serum levels of TM in patients with hypothyroidism were not significantly changed as compared with those of controls. There were a positive correlation between the serum levels of TM and FT3 as well as FT4. Serial determinations of the serum levels of TM and thyroid function(FT3, FT4 and TH) in patients with Graves' disease during treatment showed that both the serum levels of TM and thyroid hormones (FT3 and FT4) lowered progressively during treatment. After normalization of serum FT3 and FT4, the serum TM levels returned to normal. However, the serum levels of TM in patients with destructive thyroiditis and Refetoff syndrome were normal in spite of high serum levels of thyroid hormones. These data suggest that an increase in serum levels of TM is not the direct result of thyroid hormones themselves but is the result of the prolonged hypermetabolic state induced by their peripheral activities. Thyroid hormones may stimulate the synthesis or metabolism of TM on the surface of vascular endothelial cells in the patients with Graves' disease.
Graves Disease↗
[Simultaneous presence of macroamylase and macrolipase in a patient].
Here we report a case demonstrating simultaneous macroamylasemia and macrolipasemia. The patient showed persistently elevated serum amylase and lipase activities. On thin-layer gelfiltration, normal serum amylase migrated as a round spot behind albumin. However, serum amylase from this patient migrated slightly ahead of albumin. This finding indicated that serum amylase from this patient had a molecular weight greater than that of normal amylase. Lipase of normal serum was eluted from a Sephacryl S-300 column after the 4S protein. Lipase in the serum from this patient was eluted from the column slightly behind the 19S protein. This finding indicated that serum lipase from this patient also had a molecular weight greater than that of normal lipase. The conversion of macroamylase and macrolipase was observed when the normal serum was mixed with the patient's serum. To determine the isotypes of immunoglobulin (Ig) that was bound to amylase and lipase in the patient's serum, we investigated the effect of adding various anti-human Ig sera to the patient's serum. Amylase activity was precipitated only with anti-IgA lambda and lipase activity was precipitated both with anti-IgA lambda and anti-IgA kappa (anti-IgA kappa > anti-IgA lambda). Taken together, these data indicate that the material that enhanced amylase and lipase activities in the patient's serum was monoclonal IgA; IgA lambda for amylase and both IgA lambda and IgA kappa for lipase.
Amylases↗
[Present status and advances in detecting Shiga toxin-producing Escherichia coli O157].
Currently, detection of Shiga toxin-producing Escherichia coli(STEC) in stool samples is based on the isolation method in most clinical laboratories. The procedures are as follows: i) isolation with selective agar plates, ii) biological test with differential media, iii) serological test of anti-O antisera, iv) detection of toxin or toxin gene. These procedures take 4 days, therefore more rapid method is required. In the near future, a rapid detection method that detects STEC directly from stool samples will be introduced. Polymerase-chain reaction (PCR), enzyme-linked immuno-sorbent assay (ELISA), detection of serum anti-O157 antibodies are now available in clinical laboratories. Result of PCR for detection Shiga toxin gene and serum anti-O157 antibodies are described. Fifteen stool and serum samples from patients suspected of STEC infection were examined. With the culture and PCR method, 2 patients were positive by both methods and the results were confirmed in both cases. Six patients were positive by the antibodies detection method. From these results, the PCR method using stool samples was useful as a rapid detection method in clinical laboratories. Detection of serum antibodies has been simplified and is not an expensive method. Therefore, the method is useful for clinical diagnosis of STEC infection, especially, for diagnosing HUS or after antimicrobial agents have been administered to patients.
Adolescent↗
A possible involvement of Fas-Fas ligand signaling in the pathogenesis of murine autoimmune gastritis.
BACKGROUND & AIMS: A Th1 clone, II-6, established from an autoimmune gastritis BALB/c mouse that underwent thymectomy 3 days after birth, recognized a 15 mer peptide constructing the alpha subunit of H+, K(+)-adenosine triphosphatase as antigen and induced gastritis in nu/nu mice by adoptive transfer. The aim of this study was to examine the molecular mechanism of target (parietal cells) destruction in either thymectomized or II-6 cell-transferred nu/nu mice. METHODS: Expression of Fas, major histocompatibility complex class II, and intercellular adhesion molecule 1 molecules on the gastric mucosa of these mice were immunohistochemically examined. In situ DNA fragmentation in these thymectomized or nu/nu mice was tested by the terminal deoxynucleotidyl transferase-mediated digoxigenin-deoxyuridine triphosphate nick end label (TUNEL) method. Moreover, activity of II-6 cells to induce apoptosis was tested by using the 15 mer peptide-pulsed B lymphoma cells, A20.2J, as the target. RESULTS: A portion of parietal cells in gastritis-bearing thymectomized or nu/nu mice at an early stage expressed Fas, major histocompatibility complex class II, and intercellular adhesion molecule 1 molecules and was TUNEL positive. Fas-ligand message was induced on activated II-6 cells and caused DNA fragmentation of the antigen-pulsed A20.2J cells. CONCLUSIONS: Cognate interaction between Fas antigen on the target and Fas ligand on the effector seems to be one possible mechanism for the target cell destruction in organ-specific autoimmune gastritis.
Animals↗
[Development of highly sensitive assay for detection of low serum level of PIVKA-II and its clinical usefulness].
PIVKA-II is well known as a tumor marker of hepatocellular carcinoma. We describe the adaptations to render a commercially available PIVKA-II immunoassay kit (Eitest MONO P-II: Eisai Co., Ltd, Tokyo) more sensitive and evaluated clinical usefulness of this highly sensitive assay for early diagnosis of hepatocellular carcinoma. We measured serum PIVKA-II concentrations with the Eitest kit as described for the kit. To measure serum PIVKA-II concentrations below the sensitivity limit of the Eitest kit (0.0625AU/ml), linearity was determined using a series of dilutions of standard antigen diluted in standard diluent, including PIVKA-II concentrations of 0.002, 0.004, 0.008, 0.016, 0.031, 0.0625AU/ml. The assay was found to be liner from 0.004AU/ml. In this way, one can detect as low PIVKA-II concentrations as 0.004AU/ml. The repeatability-assay coefficients of variation (CV%) obtained from sixteen repeated assays of four sera were distributed between 3.63 and 18.75% and the reproducibility-assay coefficients of variation (CV%) obtained from ten repeated assays of four sera were distributed between 6.27 and 14.04%. We determined the serial changes in serum PIVKA-II levels of two patients with hepatocellular carcinoma after the resection of hepatic tumor. In these patients, elevation of serum PIVKA-II level determined by the highly sensitive assay preceded the detection of relapse by imaging diagnostic procedures. In summary, the assay system we developed has good accuracy and reproducibility for assaying low concentrations of PIVKA-II in serum and is suitable for detecting small increases in PIVKA-II concentrations. This assay system may be useful to earlier detect a relapse of hepatocellular carcinoma.
Biomarkers↗
Breakdown of self-tolerance by intrathymic injection of a T-cell line inducing autoimmune gastritis in mice.
Autoimmune gastritis (AIG) develops spontaneously in BALB/c mice thymectomized 3 days after birth (3d-Tx). We first confirmed our previous observations that CD4+ splenic T cells in AIG mice induced AIG in nu/nu mice, while those in normal mice suppressed the development of the disease. In addition, we found that a quantitative balance between these effector (Te) and suppressor (Ts) T cells determined either onset or prevention of the disease. Peripheralization of Ts seemed to begin around 3 days after birth, since the incidence of AIG in mice that underwent Tx 6 days after birth (6d-Tx) decreased markedly, compared with that of 3d-Tx mice; 12% in the former, while 79% in the latter. Notably, Ts existed in the 6d-Tx mice that escaped AIG. We next examined the target specificity of such Ts using syngeneic parietal cells known as autoantigens and two kinds of T-cell lines established from an AIG mouse; one is gastritis inducible in vivo, termed A-II, while another is not, named AC-II. Intrathymic injection of parietal cells into mice 3 days after birth followed by 6d-Tx completely prevented the development of AIG. In contrast, injection of irradiated A-II, but not AC-II cells resulted in AIG in 67% of the mice. No autoimmune oophoritis (AIO) was induced in female mice, implying that the breakdown of tolerance is organ specific. Taken together, peripheral tolerance for organ-specific autoantigens seems to be maintained by CD4+ Ts responding to Te, which induces the disease.
Animals↗
[Study for clinical usefulness of CEA, TPA, CA15-3 and BCA225 in breast cancer].
CEA, TPA, CA15-3 and BCA225 are the most frequently assayed as tumor markers for breast cancer. The aim of the present study was to reinvestigate the clinical usefulness of these markers in patients with breast cancer (primary: 28, recurrent: 15). In the cases of primary breast cancer, including all stages, positive rates of CEA, TPA, CA15-3 and BCA225 were 18%, 25%, 11% and 18%, respectively, while in the advanced cases of stage III+IV, positive rate of both CEA and TPA increased to approximately 40%. The positive rate of combination assays using four tumor markers was 46% in the cases of primary breast cancer including all stages and was 73% in the cases of advanced primary breast cancer (stage III+IV). In the cases of recurrent breast cancer, the positive rates of CEA, TPA, CA15 3 and BCA225 were 60%, 60%, 67% and 60%, respectively. When all of those tumor markers were used for assays, all of the patients with recurrent breast cancer studied showed elevated serum levels of at least one of these tumor markers. We retrospectively analyzed serum levels of tumor markers in 15 patients who relapsed breast cancer after surgical resection. In 2 patients elevation of some of four tumor markers occurred before the recurrent tumor was detected with RI scintillation and computed tomography. These results suggested that combination assays using CEA, TPA, CA15-3 and BCA225 should be helpful for detecting the recurrence of breast cancer, and also detecting the advanced primary breast cancer.
Antigens, Tumor-Associated, Carbohydrate↗
[Cytokines in laboratory medicine].
A cytokine was first recognized in 1965, and many kinds of cytokines have been found since then. Cytokines are the biologically active substances which are produced in immuno-competent cells, macrophages and monocytes, excluding immunoglobulins. They form complex networks among themselves, and mainly act locally. A single cytokine may have physiological actions to many different cells, while a certain physiological action may be shared by many different cytokines. Some of them could be now quantitated immunochemically by using highly sensitive immunoassays. Therefore, immunoassay of some cytokines may become very important in laboratory medicine.
Cytokines↗