Experimental Study of Interference in Calcitonin Testing: A Case Report and Literature Review.

BACKGROUND: Calcitonin (CT) is a sensitive serum marker of medullary thyroid carcinoma usually detected via immunoassays; however, its levels are easily disturbed by several endogenous factors. OBJECTIVE: The study aimed to discuss a case of suspected interference resulting in aberrant CT values and review previous reports of CT interference. METHODS: A female patient visited our clinic with a physical ultrasound examination showing a slightly enlarged thyroid gland with small nodules. She had elevated CT levels, inconsistent with the clinical presentation and other findings. We evaluated the results by retesting using the same platform, platform validation, multiplex dilution, Polyethylene Glycol (PEG) precipitation, heterophilic blocking tubes, and RET gene analysis. RESULTS: Retesting CT using the same platform confirmed the high value obtained. However, serial dilution of the sample produced nonlinear results, suggesting some interference. While PEG precipitation did not significantly reduce the CT level, incubating the sample in HBTs normalized the CT value, indicating interference from heterophilic antibodies. Gene sequencing revealed no RET mutations. CONCLUSION: In cases where elevated CT levels are inconsistent with clinical presentations and other findings, the laboratory technicians should communicate with clinicians, analyze the reasons for the inconsistent results, and use different methods to verify the results. Accurate testing provides realistic and reliable data for doctors and patients and helps to avoid unnecessary procedures.

Changes in Isoleucine, Sarcosine, and Dimethylglycine During OGTT as Risk Factors for Diabetes.

CONTEXT: Current metabolomics studies in diabetes have focused on the fasting state, while only a few have addressed the satiated state. OBJECTIVE: We combined the oral glucose tolerance test (OGTT) and metabolomics to examine metabolite-level changes in populations with different glucose tolerance statuses and to evaluate the potential risk of these changes for diabetes. METHODS: We grouped participants into those with normal glucose tolerance (NGT), impaired glucose regulation (IGR), and newly diagnosed type 2 diabetes (NDM). During the OGTT, serum was collected at 0, 30, 60, 120, and 180 minutes. We evaluated the changes in metabolite levels during the OGTT and compared metabolic profiles among the 3 groups. The relationship between metabolite levels during the OGTT and risk of diabetes and prediabetes was analyzed using a generalized estimating equation (GEE). The regression results were adjusted for sex, body mass index, fasting insulin levels, heart rate, smoking status, and blood pressure. RESULTS: Glucose intake altered metabolic profile and induced an increase in glycolytic intermediates and a decrease in amino acids, glycerol, ketone bodies, and triglycerides. Isoleucine levels differed between the NGT and NDM groups and between the NGT and IGR groups. Changes in sarcosine levels during the OGTT in the diabetes groups were opposite to those in glycine levels. GEE analysis revealed that during OGTT, isoleucine, sarcosine, and acetic acid levels were associated with NDM risks, and isoleucine and acetate levels with IGR risks. CONCLUSION: Metabolic profiles differ after glucose induction in individuals with different glucose tolerance statuses. Changes in metabolite levels during OGTT are potential risk factors for diabetes development.

Diagnostic performance of CA125, HE4, ROMA, and CPH-I in identifying primary ovarian cancer.

AIMS: To evaluate the ability of carbohydrate antigen 125 (CA125), human epididymis protein 4 (HE4), risk of ovarian malignancy algorithm (ROMA), and Copenhagen Index (CPH-I) to identify primary ovarian cancer (OC) from borderline and benign ovarian tumors (OTs) and explore ideal cutoff points. METHODS: A total of 684 OTs containing 276 OC patients, 116 ovarian borderline OTs and 292 benign OTs patients who underwent surgery in our hospital were included. We retrospectively searched the results of CA125 and HE4 before patients' surgery from the hospital's electronic medical records system. ROMA and CPH-I were calculated according to their menopausal status and age, respectively. Diagnostic performance of these four were assessed by drawing receiver operating characteristic (ROC) curves. RESULTS: CA125, HE4, ROMA, and CPH-I were all significantly higher in OC women compared with borderline OTs (p < 0.001), followed by benign OTs (p < 0.001). Area under the curves (AUCs) for distinguishing OC were 0.850 (0.818-0.882), 0.891 (0.865-0.916), 0.910 (0.888-0.933) and 0.906 (0.882-0.930), respectively, and the corresponding ideal cutoff values for CA125, HE4, ROMA, and CPH-I were 132.5, 68.6, 23.8, and 6.4, respectively. The difference between ROMA and CPH-I was not significant (p = 0.97), but both were higher than CA125 and HE4 (p < 0.05). HE4 showed a significantly higher AUC than CA125 (p < 0.05). For postmenopausal women, CA125 performed equivalently to ROMA (p = 0.73) and CPH-I (p = 0.91). CONCLUSIONS: In identifying patients with OC, ROMA and CPH-I outperformed single tumor marker. The diagnostic performance of HE4 was significantly higher than that of CA125. CA125 was more suitable for postmenopausal women.

A case of atypical reninoma with mild hypertension and normal plasma renin activity but elevated plasma renin concentration.

BACKGROUND: Reninoma is a rare, benign renal neoplasm. Typical clinical features include severe hypertension, secondary hyperaldosteronism, hypokalaemia and metabolic alkalosis caused by the overproduction of renin. CASE PRESENTATION: A 25-year-old lean Chinese woman with no family history of hypertension was hospitalized for stage 1 hypertension that gradually developed over two years. Endocrine investigation showed hyperreninemia without hyperaldosteronism and hypokalaemia. Interestingly, although the patient had an elevated plasma renin concentration (PRC), her plasma renin activity (PRA) was in the normal range. Abdominal contrast-enhanced computed tomography (CT) scanning revealed a solid, low-density, renal cortical mass with delayed enhancement. Selective renal vein sampling (SRVS) was performed, and a lateralization of the renin secretion from the left kidney was found. Enucleation of the tumour led to a rapid remission of hypertension and hyperreninemia. Based on pathological findings, the patient was diagnosed with reninoma. Immunohistochemical staining of the tumour was positive for Renin, CD34, Vimentin, and synaptophysin (Syn) and negative for somatostatin receptor 2 (SSTR2) and chromogranin A (CgA). CONCLUSIONS: Reninoma can present as mild hypertension without hyperaldosteronism and hypokalaemia. The clinical features of reninoma may depend on the degree of activation of the renin-angiotensin-aldosterone system (RAAS). PRC should be incorporated in the differential diagnosis of secondary hypertension.

Experimental study on abnormal thyroid function in patients with Hashimoto's thyroiditis caused by interference of thyroid hormone autoantibodies.

INTRODUCTION: Thyroid hormone autoantibody (THAAb) is one of the important factors affecting thyroid function measurement. By analyzing the examination of a patient suffered with Hashimoto's thyroiditis, we sought to find a correct assessment method. MATERIAL AND METHODS: Radioimmunoassay, chemiluminescence immunoassay on an ADVIA Centaur XP system and Architect i2000sr platform, and electrochemiluminescence immunoassay on a Roche Cobas 601 system were used for detecting thyroid function. Polyethylene glycol (PEG) precipitation were performed to eliminate the influence of THAAbs. RESULTS: The results showed that the patient's thyroid function was consistent with the clinical manifestations and conformed to the law of the hypothalamic-pituitary-thyroid axis at Architect-i2000sr platform and Roche-Cobas-601 system. The content of FT4 was significantly reduced and lower than the normal reference range, after the patients' serum was treated with PEG, which was in line with the clinical practice. The serum THAAb titer of the patients was nearly 100 times higher than that of the control group. CONCLUSIONS: Considering an abnormal thyroid function examination, it is necessary for laboratory staff to retest samples on different platforms. It is of great significance to provide a true and accurate result to clinicians and patients.

Development and validation of a novel index for the differential diagnosis of corticotropin-dependent Cushing syndrome.

PURPOSE: To develop an index for the differential diagnosis of corticotropin-dependent Cushing syndrome (CS). METHODS: The development cohort included 112 consecutive patients with clinicopathologically confirmed corticotropin-dependent CS at the Department of Endocrinology and Metabolism, Tianjin Medical University General Hospital, from December 2004 to May 2020, and data of 126 patients from studies published from 2016 to August 2020, identified through search in PubMed, Embase and the Cochrane Library, was extracted for external validation. The index was calculated as the product of plasma adrenocorticotropic hormone (ACTH, pmol/L) and urinary free cortisol (UFC, nmol/24 h) divided by 10,000. The discriminative ability was tested using receiver operating characteristics (ROC) curve analysis. RESULTS: In development cohort, area under curve of ROC analysis of the ACTH-UFC index in identifying Cushing disease (CD) was 0.977. The diagnostic accuracy of ACTH-UFC index ≤ 11 was comparable to that of 48 h 8 mg/d high-dose dexamethasone test (HDDST) in identifying CD, with sensitivity, specificity, positive and negative likelihood ratios of 96.6%, 87.5%, 7.73, and 0.04, respectively. The sensitivity of ACTH-UFC index ≤ 11 in parallel combination with pituitary magnetic resonance imaging (MRI) was 100% for identifying CD. The performance of the ACTH-UFC index in parallel or serial combination with pituitary MRI was similar in the validation cohort. CONCLUSIONS: ACTH-UFC index provides a rapid, convenient and non-invasive adjunctive approach for the differential diagnosis of corticotropin-dependent CS, with no risk of aggravating metabolic disturbances. Investigations for ectopic causes of corticotropin-dependent CS should be performed with ACTH-UFC index > 11 and negative contrasted pituitary MRI.

Clinical diagnostic performance of a fully automated TSI immunoassay vs. that of an automated anti­TSHR immunoassay for Graves' disease: a Chinese multicenter study.

BACKGROUND: Both thyroid-stimulating immunoglobulins immunoassay (TSI IA) and thyrotrophin receptor antibody immunoassay (TRAb IA) are commonly used for the diagnosis of Graves' disease (GD). The aim of the present study was to compare the clinical diagnostic performance of these two methods. METHODS: Sera were obtained from 1103 subjects presenting a variety of clinical conditions from three centers: 100 subjects with untreated GD, 200 with treated GD, 62 with autoimmune thyroid disease(AIT), 216 with other thyroid diseases (OTHER-T), 214 with non-thyroid autoimmune diseases (NTAD), 191 with other diseases (OD), and 120 healthy subjects (HS). Both TSI and TRAb IAs were performed for all 1013 serum samples. Bioassay was performed for 86 samples whose TSI results were inconsistent the TRAb assay results. RESULTS: Comparing untreated GD patients with the control groups (AIT, NTAD, OTHER-T, OD, and HS) resulted in an area under the curve (AUC) of 0.992 for the TSI IA and 0.989 for the TRAb IA with no statistically significant difference observed between these AUC values (P = 0.2733). The best TSI CDP (clinical decision point) value was 0.42 IU/L. The differences in sensitivity (100% vs. 95%, P = 0.7991) and specificity (97.1% vs. 97.6%, P = 0.9426) between the TSI and TRAb IA were not statistically significant. TSI IA had a higher agreement with the TSI bioassay than TRAb IA. CONCLUSION: The clinical diagnostic performance of the TSI IA for diagnosing Graves' disease was very similar to that of the TRAb IA. TSI IA can be used to diagnose GD in the Chinese.

Impaired Glucose-Stimulated Proinsulin Secretion Is an Early Marker of ß-Cell Impairment Before Prediabetes Stage.

CONTEXT: Evidence indicates that there is substantial impairment/loss of ß-cell function/mass even before prediabetes. Elevated plasma proinsulin is a sign of ß-cell dysfunction in patients with diabetes/prediabetes. However, the dynamic changes of glucose stimulated proinsulin secretion (GSPS) among nondiabetic individuals remain obscure. OBJECTIVE: To examine GSPS and glucose-stimulated insulin secretion (GSIS) among individuals with normal glucose tolerance (NGT) and impaired glucose tolerance (IGT) and to evaluate whether impaired GSPS is an early biomarker of ß-cell impairment in individuals with NGT who have subthreshold postprandial plasma glucose (PPG). DESIGN AND PARTICIPANTS: We evaluated GSPS and GSIS in 116 Chinese adults without diabetes (mean age ± SD, 33.31 ± 9.10 years; mean BMI, 25.24 ± 4.20 kg/m2) with fasting plasma glucose (FPG) < 5.6 mmol/L. Based on 2hPPG, the participants were divided into three groups: NGT1 (2hPPG < 6.67 mmol/L), NGT2 (6.67 ≤ 2hPPG < 7.78 mmol/L), and IGT (7.78 ≤ 2hPPG<11.1 mmol/L). We analyzed the association of GSIS and GSPS with commonly used indexes of ß-cell function, insulin resistance and family history of diabetes. RESULTS: Although not diagnosed with prediabetes, the individuals with NGT2 have clinical characteristics and high diabetes risk factors similar to those of the IGT group. However, unlike individuals with IGT, NGT2 participants did not exhibit a delayed GSIS. Instead, GSPS was impaired in NGT2 groups but not in NGT1 group. CONCLUSIONS: This study suggests that impaired GSPS, but not impaired GSIS, may serve as an early biomarker to identify a subpopulation of NGT with a high risk of diabetes.

Effect of microRNA-21 on multidrug resistance reversal in A549/DDP human lung cancer cells.

Lung cancer is a predominant cause of cancer-related mortality and numerous lung cancer patients succumb to the disease due to drug resistance. A number of microRNAs (miRNAs) are upregulated in cancer and are involved in tumorigenesis, functioning as oncogenes. Several functional studies have shown that miR-21 is important in carcinogenesis; however, none of these studies has investigated multidrug resistance (MDR) reversal in human lung cancer cells. In the present study, the effect of miR-21 on MDR reversal was analyzed in A549/DDP lung cancer cells. The data demonstrated the following after miR-21 silencing: Proliferation of the tumor cells was inhibited, cell apoptosis and oxidative damage were increased, the cell cycle was blocked at the G0/G1 phase, expression levels of P-glycoprotein were reduced, accumulation of Rhodamine 123 was increased, and the MDR-related genes encoding MDR1, MPR, glutathione S-transferase-π, B-cell lymphoma 2, cyclin-dependent kinase 1, cystathione and glutathione were downregulated. Further mechanistic analysis revealed that miR-21 silencing reduced AKT phosphorylation and transcriptional activation of E2F-1 and Twist. In conclusion, this study demonstrated that miR-21 silencing reversed lung cancer cell MDR by modulation of MDR-related gene expression and inhibition of the AKT signaling pathway, suggesting that miR-21 may be a potential therapeutic candidate in patients with MDR lung cancer.

Bone metastasis from breast cancer involves elevated IL-11 expression and the gp130/STAT3 pathway.

To evaluate the relationship between IL-11 and bone metastasis in patients with breast cancer and explore the potential molecular mechanism, total serum samples were collected from 180 breast cancer patients and 20 women without breast cancer. The serum expression level of interleukin (IL)-11, connective tissue growth factor (CTGF), transforming growth factor-ß, and Tracp5b was determined by enzyme-linked immunosorbent assay, and mRNA expression of IL-11 in fresh breast cancer tissue was determined by RT-PCR. Immunohistochemical staining was used to detect the expression of IL-11 and CTGF in breast cancer tissue, and Western blot was used to detect the expression of p-38, p-C-JUN, p-STAT3, and p-gp130 in fresh breast cancer tissue. DNA-binding activity of AP-1 was examined by electrophoretic mobility shift assay. Differences were statistically analyzed between the group with breast cancer metastatic to bone (MBC-B) and the group with only primary breast cancer (PBC). Serum level and mRNA expression of IL-11 in the MBC-B group were significantly higher than those in the PBC group. IL-11 immunohistochemical staining showed that the percentage of positively stained cells in the MBC-B group (57.5 %) was significantly higher than that in the PBC group (14.29 %). Western blot analysis showed higher expression of p-p38, p-C-JUN, p-STAT3, and p-gp130 in the MBC-B group than in the PBC group. DNA-binding activity of AP-1 was significantly higher in the MBC-B group than in the PBC group. These data suggest that IL-11 is associated with bone metastasis and may be of value for predicting bone metastasis from breast cancer.

Trimester- and method-specific reference intervals for thyroid tests in pregnant Chinese women: methodology, euthyroid definition and iodine status can influence the setting of reference intervals.

OBJECTIVE: The importance of diagnosis and treatment of thyroid dysfunction during pregnancy has been widely recognized. We therefore established trimester- and method-specific reference intervals for thyroid testing in pregnant women according to the NACB recommended criteria. Several factors can affect the setting of reference intervals, in particular manufacturer's methodology, euthyroid definition and iodine status. DESIGN: Cross-sectional dataset analysis. SUBJECTS: Five hundred and five normal pregnant women at different stages of gestation were rigorously selected for setting reference intervals. All were healthy, iodine sufficient, euthyroid and negative for both serum thyroid peroxidase antibody (TPOAb) and thyroglobulin antibody (TgAb). MEASUREMENTS: Thyrotrophin (TSH), total and free thyroxine (TT4 and FT4), total and free triiodothyronine (TT3 and FT3) and anti-TPOAb and anti-TgAb were measured using the Bayer ADVIA Centaur system. Iodine content in drinking water, salt and urine was determined by national standard methods. The 2·5th and 97·5th percentiles were calculated as the reference intervals for thyroid hormone levels during each trimester. RESULTS: All participants had long-term consumption of iodized salt and median urinary iodine of 150-200 µg/l during each three trimester. The reference intervals for the first, second and third trimesters were, respectively, TSH 0·03-4·51, 0·05-4·50 and 0·47-4·54 mIU/l and FT4 11·8-21·0, 10·6-17·6 and 9·2-16·7 pmol/l. The manufacturer's method, euthyroid definition and iodine status may influence TSH and FT4 reference intervals. Alterations in thyroid hormone concentrations during pregnancy differed at different stage of gestation and to those of a nonpregnant state. CONCLUSIONS: The trimester- and method-based reference intervals for thyroid tests during pregnancy are clinically appropriate. Some variables should be controlled when establishing reference intervals.

Signal pathways involved in emodin-induced contraction of smooth muscle cells from rat colon.

AIM: To investigate the effects induced by emodin on single smooth muscle cells from rat colon in vitro, and to determine the signal pathways involved. METHODS: Cells were isolated from the muscle layers of Wistar rat colon by enzymatic digestion. Cell length was measured by computerized image micrometry. Intracellular Ca2+ ([Ca2+]i) signals were studied using the fluorescent Ca2+ indicator fluo-3 and confocal microscopy. PKCalpha distribution at rest state or after stimulation was measured with immunofluorescence confocal microscopy. RESULTS: (1) Emodin dose-dependently caused colonic smooth muscle cells contraction; (2) emodin induced an increase in intracellular Ca2+ concentration; (3) the contractile responses induced by emodin were respectively inhibited by preincubation of the cells with ML-7 (an inhibitor of MLCK) and calphostin C (an inhibitor of PKC); (4) Incubation of cells with emodin caused translocation of PKCalpha from cytosolic area to the membrane. CONCLUSION: Emodin has a direct contractile effect on colonic smooth muscle cell. This signal cascade induced by emodin is initiated by increased [Ca2+]i and PKCalpha translocation, which in turn lead to the activation of MLCK and the suppression of MLCP. Both of them contribute to the emodin-induced contraction.

Prospective study of biofeedback retraining in patients with chronic idiopathic functional constipation.

AIM: To determine the efficacy and long-term outcome of biofeedback treatment for chronic idiopathic constipation and to compare the efficacy of two modes of biofeedback (EMG-based and manometry-based biofeedback). METHODS: Fifty consecutive contactable patients included 8 cases of slow transit constipation, 36 cases of anorectic outlet obstruction and 6 cases of mixed constipation. Two modes of biofeedback were used for these 50 patients, 30 of whom had EMG-based biofeedback, and 20 had manometry-based biofeedback. Before treatment, a consultation and physical examination were done for all the patients, related information such as bowel function and gut transit time was documented, psychological test (symptom checklist 90, SCL90) and anorectic physiological test and defecography were applied. After biofeedback management, all the patients were followed up. The Student's t-test, chi-squared test and Logistic regression were used for statistical analysis. RESULTS: The period of following up ranged from 12 to 24 months (Median 18 months). 70 % of patients felt that biofeedback was helpful, and 62.5 % of patients with constipation were improved. Clinical manifestations including straining, abdominal pain, bloating, were relieved, and less oral laxative was used. Spontaneous bowel frequency and psychological state were improved significantly after treatment. Patients with slow and normal transit, and those with and without paradoxical contraction of the anal sphincter on straining, benefited equally from the treatment. The psychological status rather than anorectal test could predict outcome. The efficacy of the two modes of biofeedback was similar without side effects. CONCLUSION: This study suggests that biofeedback has a long-term effect with no side effects, for the majority of patients with chronic idiopathic constipation unresponsive to traditional treatment. Pelvic floor abnormalities and transit time should not be the selection criteria for treatment.

Contractile effects and intracellular Ca2+ signalling induced by emodin in circular smooth muscle cells of rat colon.

AIM: To investigate whether emodin has any effects on circular smooth muscle cells of rat colon and to examine the mechanism underlying its effect. METHODS: Smooth muscle cells were isolated from the circular muscle layer of Wistar rat colon and the cell length was measured by computerized image micrometry. Intracellular Ca(2+) ([Ca(2+)]i) signalling was studied in smooth muscle cells using Ca(2+) indicator Fluo-3 AM on a laser-scanning confocal microscope. RESULTS: Emodin dose-dependently induced smooth muscle cells contraction. The contractile responses induced by emodin were inhibited by preincubation of the cells with ML-7 (an inhibitor of MLCK). Emodin caused a large, transient increase in [Ca(2+)]i followed by a sustained elevation in [Ca(2+)]i. The emodin -induced increase in [Ca(2+)]i was unaffected by nifedipine, a voltage-gated Ca(2+)-channel antagonist, and the sustained phase of the rising of [Ca(2+)]i was attenuated by extracellular Ca(2+) removal with EGTA solution. Inhibiting Ca(2+) release from ryanodine-sensitive intracellular stores by ryanodine reduced the peak increase in [Ca(2+)]i. Using heparin, an antagonist of IP(3)R, almost abolished the peak increase in [Ca(2+)]i. CONCLUSION: Emodin has a direct excitatory effect on circular smooth muscle cells in rat colon mediated via Ca(2+)/CaM dependent pathways. Furthermore, emodin-induced peak [Ca(2+)]i increase may be attributable to the Ca(2+) release from IP(3) sensitive stores, which further promote Ca(2+) release from ryanodine-sensitive stores through CICR mechanism. Additionally, Ca(2+) influx from extracellular medium contributes to the sustained increase in [Ca(2+)]i.

Etiology and management of hemmorrhage in spontaneous liver rupture: a report of 70 cases.

AIM: To analyze the causes and management of hemorrhage in spontaneous liver rupture. METHODS: Seventy cases of spontaneous liver rupture were retrospectively analyzed for causes of hemorrhage and therapeutic effects of surgical approaches. RESULTS: It was demonstrated that the causes of spontaneous liver rupture were primary liver cancer in 60 cases (85.7 %), cirrhosis in 3 cases (4.3 %), liver angioma in 2 cases (2.9 %), liver adenoma in 4 cases (5.7 %),and secondary liver cancer in 1 case (1.4 %). Hemostasis was achieved with surgical approaches in 68 cases (97.1 %) and non-surgical approaches in 2 cases (2.9 %). Surgical interventions included suture, ligation of hepatic artery, hepatic artery chemoembolization and partial hepatic resection. CONCLUSION: The results suggest that surgical intervention is still the main therapeutic method and the best procedure that should be selected according to causes of disease and patient's condition and history.