Assessment of humoral responses in COVID-19 using various quantitative antibody tests.

BACKGROUND: Quantitative antibody tests are expected to be useful in diagnostics of COVID-19 and investigation of herd immunity against SARS-CoV-2. To make it proper to perform them, understanding of the immunological aspects is critically important. The present study aimed to assess humoral responses in COVID-19 using various quantitative antibody tests. METHODS: Four quantitative antibody tests that are different in targeted antigens, detectable immunoglobulin classes and avidity were used. Diagnosis was confirmed by RT-PCR for SARS-CoV-2 detection. Antibody titres of 117 samples collected from 24 COVID-19 patients and 23 non-COVID-19 patients were measured to evaluate correlations between different tests. For 24 COVID-19 patients, antibody titres measured at various time points after the onset or the RT-PCR diagnosis were subjected to assessment of humoral responses. RESULTS: Correlations between tests were observed to some degree, although there were discrepancies putatively due to differences in measurement principle. Seronegative COVID-19 was diagnosed for some patients, in whom antibody titres were less than the cut-off value in each test throughout the time courses. IgG seroconversion without prior IgM seroconversion most frequently occurred, while predominance of IgM responses over IgG responses was observed in some severe cases. Viral burdens estimated according to threshold cycle values at the RT-PCR seemed to impact antibody responses. CONCLUSIONS: The results provide insights into the nature of humoral responses to SARS-CoV-2 and diagnostic performance of antibody tests.

[Post-graduate Education of Clinical Laboratory Physicians and Medical Technologists (Biomedical Laboratory Scientist) -Chairpersons' Introductory Remarks -].

The symposium was cosponsored by the College of Laboratory Medicine of Japan (CLMJ). Two main top- ics were discussed in it. One was for clinical laboratory physicians, and the other was for medical technolo- gists. Regarding the former theme, a lecture was held about the medical specialist system beginning in the 2017 fiscal year, and the educational steps for becoming a clinical laboratory physician in the system. Re- garding the latter, four lectures were held, 1. Post-graduate education for medical technologists and the role of the Japanese Association of Medical Technologists, 2. Certification examinations by CJMJ, 3. Certification examinations by the College of Molecular Analysis, 4. Certification systems in foreign countries. We hope that this symposium can promote understanding of post-graduate education for clinical laboratory physicians and medical technologists, leading to career improvement. [Review].

[Present Status and Problems of Laboratory Test in a Community and at Home: "Sample Measurement Room", Self-Test and Mail-in Test.]

In this review, the present status and problems of laboratory tests outside traditional healthcare were dis- cussed. The test group includes the "sample measurement room", self-test, and mail-in test. They will be classified by 2 X 2 viewpoints, sampling/measurement X place/person. As for the "sample measurement room", consultation based on the results cannot be recommended, even if the results are abnormal, although this is often misunderstood. The problems of the "sample measurement room", self-test, and mail-in test can be considered from the viewpoints mentioned above. A sample extraction defect, infection between persons, the data instability by preservation, measurement error, and insufficient quality control are thought to be examples.

[Quality assurance of point of care testing].

Quality assurance of point of care testing (POCT) is thought to be difficult since it is mainly performed by doctors or nurses away from the laboratory. According to POCT guideline second edition, published by The Japan Society for Clinical Laboratory Automation, all the stages of POCT, i.e. from "before sample collection" to "after result reporting", need to be managed. In these stages, problems in the management of reagents, the quality control of measurements, and preservation of test results in medical records in POCT were discussed. Although there are problems, progress in equipment and reagents to assure POCT has been accomplished. Considering the POC coordinator's role, many activities are associated with quality assurance and are indispensable; however, as all laboratory staff can coordinate POCT, information about what is needed at the point of care and should be collected put into action. Our laboratory's participation in blood glucose measurements on the ward and in outpatients is described as an example.

A case of atypical POEMS syndrome without polyneuropathy.

POEMS (Polyneuropathy, Organomegaly, Endocrinopathy, M-protein, Skin changes) syndrome is a rare hematological disease associated with overproduction of pro-inflammatory cytokines. Under the current nomenclature and diagnostic criteria for POEMS syndrome, the presence of characteristic polyneuropathy is required for diagnosis. We report a 43-year-old Japanese woman with organomegaly, endocrinopathy, M-protein, skin lesions, as well as typical renal lesions and sclerotic bone lesions. Of note, neurological examinations and peripheral nerve conduction tests were normal in this patient. In view of the overwhelming number of otherwise characteristic signs and symptoms, we made a provisional diagnosis of 'atypical POEMS syndrome without polyneuropathy'. If further similar cases are reported in the future, reconsideration of the nomenclature and/or diagnostic criteria for POEMS syndrome may be required.

Serum alcohol dehydrogenase: a sensitive biomarker of ongoing graft function after liver transplantation.

Alcohol dehydrogenase (ADH) is an enzyme specifically located in the cytoplasm of hepatocytes. The purpose of this study was to assess the potential usefulness of serum ADH activity as a biomarker of graft function following liver transplantation. Blood samples were obtained from 26 patients who underwent living-donor liver transplantation. In patients without any post-operative complication, serum ADH activity normalized at 2.9 +/- 1.2 d. Values of serum ADH activity were remarkably elevated in patients with vascular complications, whereas they were only slightly elevated or remained within the reference range in patients with acute cellular rejections. In vascular complications, serum ADH activity peaked prior to elevation of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), and once the cause of damage was resolved, the values returned to reference range more quickly than did ALT and AST. In conclusion, monitoring serum ADH activity in addition to ALT and AST may provide more sensitive ongoing graft status and valuable information for the differential diagnosis of vascular complications and acute cellular rejection.

[Apolipoprotein E deficiency associated with type III hyperlipoproteinemia--analysis of apolipoprotein E].

CASE REPORT: The patient was a 50-year old male who was found to have a high cholesterol level during a routine health check up at work 5 years before and was examined at Keio University Hospital. Lipoprotein electrophoresis on agarose gel revealed type III hyperlipidemia, and a screening test yielded the following values (mg/dl): total cholesterol, 420; TG, 138; and HDL-cholesterol, 105. Turbidimetric immunoassay showed that the apolipoprotein E (apoE) level was below the limit of detection. Since he was 25 years old, the patient had sometimes noticed xanthomas on his knees and eyelids, and for that reason we made a diagnosis of apoE deficiency associated with type III hyperlipidemia. We tried using SDS-polyacrylamide gel electrophoresis, Western blot, and the protein chip method to detect apoE in this case, but the level was below the limit of detection by the first two methods, and it was so low that it was detected near the sensitivity limit of the protein chip method. Diet therapy, statin therapy, and fibrate therapy have been continued, and the latest data are: total cholesterol, 373; TG, 95; and HDL-cholesterol, 83. No manifestations associated with arteriosclerotic disease other than mild xanthomas have been observed.

Salt intake affects the relation between hypertension and the T-786C polymorphism in the endothelial nitric oxide synthase gene.

BACKGROUND: Recent genetic studies have shown an association between the T-786C polymorphism in the endothelial NO synthase gene and coronary artery diseases, but any possible association with hypertension has been controversial. Other studies indicate the effect of restricting salt intake differ depending on individual salt-sensitivity, and the mixture of different sensitivity in study subjects may obscure the results. The objective of this study was to investigate the gene-environment interaction between the salt intake and this polymorphism. METHODS: We genotyped 281 healthy men after excluding 37 men on hypertensive therapy (mean age 44.8 +/- 11.9 years) for the mutation, and evaluated their daily salt intake using a validated food frequency questionnaire. RESULTS: A quartile classification of salt intake revealed that the blood pressure of subjects with the mutation was significantly higher than that of subjects without the mutation, but only in the 4th quartile (the highest intake group). A multiple logistic regression analysis also showed that the presence of this mutation increased the risk of hypertension only in the 4th quartile (adjusted odds ratio = 6.38, P = .025). CONCLUSIONS: The presence of this mutation alone does not significantly increase the risk of hypertension. However, high salt intake interacts with the mutation and leads to a significant increase in the risk of hypertension. The T-786C mutation warrants being considered a candidate for further study with the aim of tailor-made hypertension prevention.

Increased risk of obesity resulting from the interaction between high energy intake and the Trp64Arg polymorphism of the beta3-adrenergic receptor gene in healthy Japanese men.

BACKGROUND: Few studies have investigated the interaction between the Trp64Arg polymorphism of the beta3-adrenergic receptor gene (ADRB3) and environmental factors. This study aimed to investigate whether energy intake affects the relationship between this polymorphism and obesity. METHODS: Healthy Japanese men (n=295; age 46.1+/-11.5 years (mean +/-standard deviation); waist circumference 83.9+/-9.3 cm; body mass index (BMI) 23.3+/-3.3 kg/m2) recruited in a Japanese chemical industry firm were eligible for analysis. Daily energy intake, protein, fat, and carbohydrate (PFC) ratio and daily physical activity were assessed by self-reported questionnaires. Genotyping for the polymorphism was performed with written informed consent. RESULTS: When the subjects were classified into two groups according to presence of the polymorphism, the groups were not significantly different in waist circumference or BMI. Quartile classification of energy intake, however, demonstrated a significantly larger ratio of obese subjects to non-obese subjects in the group with the polymorphism in the highest 4th quartile alone. Multiple logistic regression analysis also revealed that the presence of the polymorphism increased the risk of obesity significantly in the 4th quartile alone (adjusted odds ratio=3.37, 95% confidence interval=1.12-10.2). CONCLUSION: Presence of the polymorphism alone does not significantly increase the risk of obesity. However, high energy intake interacts with the polymorphism and leads to a significant increase in risk of obesity. The Trp64Arg polymorphism of ADRB3 warrants consideration, along with other polymorphisms involved in the development of obesity, for tailor-made prevention of obesity.

Off-pump coronary artery bypass attenuates transient hepatocellular damage after myocardial revascularization.

OBJECTIVE: Cardiopulmonary bypass (CPB) affects hepatocellular integrity and occasionally results in liver dysfunction after cardiac surgery. Performing coronary artery bypass graft surgery without CPB may help to reduce the risk of this complication and better preserve perioperative liver function. This study compared perioperative hepatocellular damage in patients undergoing on-pump and off-pump bypass surgery. DESIGN: Prospective study. SETTING: University hospital. PARTICIPANTS: Patients scheduled for elective on-pump (n = 21) and off-pump (n = 17) coronary artery bypass surgery. MEASUREMENTS AND MAIN RESULTS: Liver function was assessed by serum levels of alcohol dehydrogenase (AD) and alpha-glutathione S-transferase (alpha-GST), which serve as more sensitive indices of hepatocellular injury than do conventional transaminases. Arterial blood was sampled at 6 stages: after induction of anesthesia (baseline); at the end of CPB in the on-pump group or on completion of the last distal anastomosis in the off-pump group; at the end of surgery; and 6 hours, 12 hours, and 24 hours after the end of anesthesia. The off-pump patients showed significantly lower increases in serum AD and alpha-GST levels than did the on-pump group. AD and alpha-GST values increased in the on-pump patients after the initiation of CPB and peaked at the end of surgery, with a return to baseline at 12 hours and 24 hours after the end of anesthesia. No clinically relevant liver dysfunction was observed in either group. CONCLUSIONS: CPB induced transient subclinical hepatocellular damage, whereas off-pump revascularization attenuated this damage.

[Analysis of small dense LDL in patients with type 2 diabetic mellitus by the modified Krauss method using an internal standard].

In patients with Type 2 diabetes mellitus (Type 2 DM), the relationship between the prevalence rate of small dense LDL (sdLDL) and parameters of lipid metabolism was analyzed using the method devised by modified Krauss method using apoferritin as an internal standard. The prevalence rate of sdLDL was 34% compared with it of normal subjects in this study. When the severity of Type 2 DM was classified into three groups of the HbA1c value, neither the sdLDL size nor its prevalence rate differed significantly depending upon the severity of the Type 2 DM. Also, when the prevalence rate of sdLDL was analyzed in relation to the severity of complications, i.e., of microangiopathy (retinopathy and nephropathy) or macroangiopathy (cerebral infarction), there was no significant difference in the prevalence rate of sdLDL depending on the severity of any of these complications. On the other hand, the prevalence rate of sdLDL was found to be correlated with the serum TG level. The serum level of TG-rich remnants (metabolites of TG) was also high in patients with sdLDL. It should take notice that the assessment of sdLDL should be used the authorized method for the evaluation. Thus it is concluded that the levels of sdLDL were important in evaluation of Type 2 DM. The prevalence rate of sdLDL did not correlate with the severity, nor the modalities for the complications of Type 2 DM.

Assessment of tailor-made prevention of atherosclerosis with folic acid supplementation: randomized, double-blind, placebo-controlled trials in each MTHFR C677T genotype.

This study aimed at assessing the effect of folic acid supplementation quantitatively in each MTHFR C677T genotype and considered the efficiency of tailor-made prevention of atherosclerosis. Study design was genotype-stratified, randomized, double-blind, placebo-controlled trials. The setting was a Japanese company in the chemical industry. Subjects were 203 healthy men after exclusion of those who took folic acid or drugs known to effect folic acid metabolism. Intervention was folic acid 1 mg/day p.o. for 3 months. The primary endpoint was plasma total homocysteine level (tHcy). In all three genotypes, there were significant tHcy decreases. The greatest decrease was in the TT homozygote [6.61 (3.47-9.76) micromol/l] compared with other genotypes [CC: 2.59 (1.81-3.36), CT: 2.64 (2.16-3.13)], and there was a significant trend between the mutated allele number and the decrease. The tHcy were significantly lowered in all the genotypes, but the amount of the decrease differed significantly in each genotype, which was observed at both 1 and 3 months. Using these time-series data, the largest benefit obtained by the TT homozygote was appraised as 2.4 times compared with the CC homozygote. Taking into account the high allele frequency of this SNP, this quantitative assessment should be useful when considering tailor-made prevention of atherosclerosis with folic acid.

[Consulting work performed by the hospital laboratory. The usefulness of the Laboratory Information Office in Keio University Hospital].

To establish a consultation office for handling questions from physicians and other hospital staff is one way to provide laboratory information for effective utilization of laboratory data. Here I introduce the basis of our Laboratory Information Office in Keio University Hospital. A full-time specialized, technologist is stationed in the office. She answers inquiries using a computer database, informs physicians of abnormal laboratory findings and maintains information exhibited on the web site of the clinical laboratory in the intranet of our hospital. The number of consultations is about 280 per month. The most common area of inquiry that of immuno-serology. Cooperation between the technologist and the clinical pathologists makes it possible to respond more efficiently to the inquiries, and in the future could lead to more useful consultations with clinical physicians. In the future, mutual communication between laboratory consultation offices could be arranged to share knowledge. Also, direct consultation with patients concerning laboratory examinations should be considered, though this may be difficult to implement.

[Tumor markers of urinary tract carcinoma].

The tumor markers for malignant tumors arisen from urinary system including prostate cancer were reviewed. As for renal cell carcinoma there was no good marker used in routine test level at present. In the diagnosis of urothelial (transitional cell) carcinoma, mainly bladder cancer, 3 methods (urinary BTA, NMP22 and BFP) are used now in Japan. They all seem to be not fully sufficient in respect of the specificity. In foreign countries, new tests such as urinary telomerase and BLCA-4 are used and have been evaluated. On the diagnosis of prostate cancer, serum total PSA is well established and used. Various PSA relation markers have been advocated for the differentiation between benign prostate hypertrophy and carcinoma in so called "gray zone" level of total PSA. In methods based on the molecular forms of PSA, the ratio of free PSA to total PSA (f/T) is widely use, and proPSA is a test that is expected. Other approaches such as volume of index PSA, age specific PSA reference range and PSA velocity are also in practical application. Human glandular kallikrein 2, which belong to the human kallikrein family as well as PSA, is expected as a tumor specific marker.

[Effective cooperation between clinical physicians and laboratory consultation division].

With the increase in clinical examination items, and with the specialization of clinical medicine, physicians' requests for consultations on laboratory medicine is getting more evident. However, at present in many hospitals laboratory staff do not always make appropriate answers to the questions from physicians in various fields because the laboratories are divided in terms of examination fields. In our hospital, we established a consultation division responding to every inquiry from clinical staff with a full-time technologist. At present the division is functioning very well, suggesting that laboratory technologists must build close relations with clinical staff and that the technologist's task is very diverse, not confined to just analyzing samples.

Alport syndrome-like basement membrane changes in Frasier syndrome: an electron microscopy study.

Frasier syndrome (FS) is a rare disease characterized by male pseudohermaphroditism and slowly progressing nephropathy. FS originates from heterozygous mutation in the intron 9 splicing donor site of Wilms' tumor suppressor gene (WT1). Focal segmental glomerular sclerosis is common in FS, but there have not been so many detailed pathologic investigations. The authors examined the kidneys of 3 patients with FS. The results showed that nephropathy started as mesangial proliferative glomerulonephritis, and later a concomitant focal segmental lesion developed. In all cases, electron microscopy results showed widespread thinning, splitting, and lamellation of the glomerular basement membrane, which mimicked hereditary nephritis. Throughout adulthood, WT1 protein expresses on glomerular podocytes. Recent reports described that podocytes expressing WT1 play an important role in maintaining the glomerular basement membrane. Hereditary nephritis-like glomerular basement membrane findings in FS suggest that one of the important functions of podocytes is to form and maintain the glomerular basement membrane.