OBJECTIVE

To evaluate the diagnostic value of CK-MB in acute myocardial infarction, to differentiate the relation between creatine phosphokinase (CPK), CK-MB and macro CK-BB (a macromolecule formed by an atypic CK-BB linked to an immunoglobulin) and to investigate the association of macro-BB with cancer.

METHODS

We studied 105 patients: 47 with various tumors; 8 with acute myocardial infarction and one control group with 50 persons without cardiac or oncologyc diseases. The CK-MB, CPK and macro CK-BB were measured in the serum of all patients and of the control group.

RESULTS

Among patients with cancer, 12 (25.6%) showed positive serum macro CK-MB. Two of them (16.6%) showed concomitant increase in CK-MB. All patients with increased serum macro CK-MB had malignant tumors.

CONCLUSIONS

The CK-MB is a reliable test for the myocardial infarction, however, when this isozyme is increased in the presence of normal CPK value, it must be questioned the presence of macro CK-BB to avoid false positive results. The presence of macro CK-BB could be relationed with malignant disease, specially with carcinoma of the prostate.

In the acute stage of head injury biochemical markers are valuable in the assessment of tissue damage and outcome. In this study, in addition to evaluating computed tomographic (CT) scans, we measured plasma aldosterone and brain type of serum creatine kinase isozyme (CK-BB) to determine whether or not the values obtained reflect the degree of morphological brain changes caused by injury. All 152 patients studied were diagnosed with CT and blood samples were taken on admission. On the basis of clinical and CT findings they were classified into seven groups according to the nature and severity of the insult. Slightly over one half had severe brain injury. Aldosterone was measured by radioimmunoassay and CK-BB by electrophoresis. Aldosterone and CK-BB levels were high in patients with severe cerebral contusion and in those whose CT scan showed a marked midline shift and disappearance of the perimesencephalic cistern. Aldosterone, but not CK-BB, was markedly elevated in patients with epidural hematoma and was only moderately increased in those with a slight midline shift. The results of this study indicate that aldosterone and CK-BB may be useful as biochemical markers in the acute stage of head injury.

Adenylate kinase (AK) activity in the cerebrospinal fluid (CSF), described as a marker of brain edema and lesions in adults, was studied in 79 newborns with severe respiratory distress within 24 h after admission to the Intensive Care Unit (ICU). The CSF-AK activity was compared with CSF lactate concentration, CSF lactate dehydrogenase activity (LDH), and CSF and serum creatine kinase isoenzyme BB (CK-BB) activity. Newborns were divided into Group I with moderate to severe brain dysfunction and Group II with mild or no detectable brain dysfunction on discharge from the ICU. Mean CSF-AK activity (11.31 U/L) in Group I was significantly (p less than 0.001) higher than in Group II (2.82 U/L). Correlation between CSF-AK and CSF lactate was r = 0.714, p less than 0.01 and between CSF-AK activity and CSF-LDH activity was r = 0.550, p less than 0.01 in Group I. Preliminary data indicate that CSF-AK activity within 24 h after ischaemia is an indicator of hypoxic brain lesions in newborns. Its prognostic value for the infant's development remains to be determined by further study.

OBJECTIVE

To report a case of chronically elevated creatine kinase (CK) concentration that is possibly associated with renal insufficiency and prostatic carcinoma. The goal is to raise awareness among clinicians who monitor CK concentrations in patients receiving hydroxymethylglutaryl-coenzyme A (HMG-CoA) reductase inhibitors.

METHODS

Because of an elevated CK concentration, a 64-year-old African-American man with a history of chronic heart disease and renal insufficiency was assessed for possible myositis relating to his treatment with HMG-CoA reductase inhibitors. However, an association between the elevated enzyme concentration and drug treatment could not be clearly established. The patient was subsequently diagnosed with prostatic cancer and underwent a radical retropubic prostatectomy. The CK enzyme concentration declined following the surgery despite continuation of the drug therapy.

CONCLUSIONS

CK is relatively nonspecific because of its wide distribution in human tissues. Although several findings of elevated CK concentrations, particularly the CK-BB isoenzyme, in patients with carcinoma or chronic renal insufficiency have been documented, these may not be common knowledge among clinicians. This case report provides an example of an unusually high CK enzyme concentration that may be linked to prostatic carcinoma and renal insufficiency.

CONCLUSIONS

It is important to be aware of different causes for CK enzyme concentration elevation, especially when it is used as a monitoring parameter during HMG-CoA reductase inhibitor treatment. In a case of persistent elevated CK enzyme concentration without evidence of myositis, renal insufficiency may be a contributing factor and malignancy must be ruled out.

We assessed the effects of angiotensin-converting enzyme (ACE) inhibition on changes in the myocardial intracellular creatine kinase (CK) system in relation to left ventricular (LV) remodeling and function in heart failure after myocardial infarction (MI) in rats. We compared the findings at 4 weeks after MI to those at 12 weeks after MI. LV weight and chamber size were significantly increased and percent fractional shortening (%FS) was decreased in untreated MI rats compared with normal control animals both at 4 and 12 weeks after MI. Animals with MI and treated with the ACE inhibitor temocapril showed significantly reduced LV weight and chamber size and increased %FS compared with untreated MI rats at 12 weeks after MI, but not at 4 weeks after MI. At 4 weeks after MI, no significant changes were found in the total creatine and relative distribution of each CK isoenzyme in either the temocapril-treated or untreated animals with MI compared with the normal controls. In contrast, at 12 weeks after MI, untreated MI rats showed significant reductions in the total creatine and mitochondrial and MM-CK fractions and increases in the MB- and BB-CK fractions compared with the controls. The alterations in the mitochondrial and MB-CK fractions were significantly attenuated after 12 weeks of ACE inhibition. Thus, LV myocardial energy metabolism is progressively impaired and its alteration is not related to the magnitude of geometric changes and LV dysfunction after MI. Most of the beneficial effects of ACE inhibition were observed at 12 weeks after MI. Our results may provide an insight into the therapeutic strategy of ACE inhibition in chronic heart failure after MI.

The effects of prostaglandin E1 (PGE1) on hepatic sinusoidal endothelial cells (SEC) in the xenogeneic immunoreaction were investigated. Porcine livers were perfused with fresh human blood via the portal vein (PV) and the hepatic artery (HA) either with the administration of PGE1 (Group PG) or without PGE1 (Group C). The creatine kinase-BB component (CK-BB) in the perfusate was measured to assess SEC damage. SEC activation and complement activation were evaluated immunohistochemically by the expression of von Willebrand factor (vWF) and by the deposition of membrane attack complex (MAC), respectively. Xenoperfusion in Group C was discontinued between 4 and 6 hr due to the rapid elevation of HA pressures and the massive loss of perfusate. In Group PG, both PV and HA pressures were kept stable for up to 9 hr. In Group C, severe interlobular bleeding and diffuse extrasinusoidal hemorrhage were observed at 4 hr histologically, while in Group PG, the hepatic architecture was maintained without hemorrhage at 6 hr. MAC was markedly deposited on SEC and parenchymal cells at 3 hr in both groups. The amount of vWF, however, was expressed on SEC in large amounts at 1 hr in Group C, while small amounts were expressed at 1 hr in Group PG. In Group PG, CK-BB release was significantly lower than in Group C (P < 0.01). These results suggest that PGE1 suppressed SEC activation and protected the impairment of hepatic SEC during xenoperfusion without suppressing complement activation, resulting in the prolongation of xenogeneic liver perfusion.

The inclusion of EDTA in the creatine kinase reagent recommended by the Scandinavian Committee on Enzymes was shown to increase reagent stability from less than 24 h to 5 days. Part of this effect can be explained by the fact that EDTA delays the formation of inhibitory products formed when N-acetyl cysteine is oxidized. The addition of EDTA to the reagent also results in increased measured CK activity. This effect is more pronounced for CK-BB than for CK-MM. Calcium and ferric ions are shown to inhibit the enzyme and the chelation of these ions can partly explain the observed increase of CK acitivity.

Increasing evidence indicates that the enzyme creatine kinase (CK) is intimately involved in microvascular contractility. The mitochondrial isoenzyme catalyses phosphocreatine synthesis from ATP, while cytoplasmic CK, predominantly the BB isoenzyme in vascular tissue, is tightly bound near myosin ATPase, where it favours ATP production from phosphocreatine to metabolically support vascular contractility. However, the effect of CK gene inactivation on microvascular function is hitherto unknown. We studied functional and structural parameters of mesenteric resistance arteries isolated from 5 adult male mice lacking cytoplasmic BB-CK and ubiquitous mitochondrial CK (CK-/-) vs 6 sex/age-matched controls. Using a Mulvany Halpern myograph, we assessed the acute maximum contractile force with 125 mM K⁺ and 10^-5 M norepinephrine, and the effect of two inhibitors, dinitrofluorobenzene, which inhibits phosphotransfer enzymes (0.1 μM), and the specific adenylate kinase inhibitor P1, P5-di(adenosine 5') pentaphosphate (10^-6 to 10^-5 M). WT and CK-/- did not significantly differ in media thickness, vascular elasticity parameters, or acute maximum contractile force. CK-/- arteries displayed greater reduction in contractility after dinitrofluorobenzene 38%; vs 14% in WT; and after AK inhibition, 14% vs 5.5% in WT, and displayed abnormal mitochondria, with a partial loss of the inner membrane. Thus, CK-/- mice display a surprisingly mild phenotype in vascular dysfunction. However, the mitochondrial abnormalities and greater effect of inhibitors on contractility may reflect a compromised energy metabolism. In CK-/- mice, compensatory mechanisms salvage energy metabolism, as described for other CK knock-out models.

Keywords: BB-creatine kinase isoenzyme; Creatine kinase; Knockout mice; Mitochondrial creatine kinase isoenzyme; Resistance arteries; Wire myograph.

Celiptium (Ce) is an antitumor drug used in the therapy of breast carcinomas, which are to a large extent dependent on estrogens. We have studied the effect of Ce on some proteins induced by estradiol (E2) in the rat uterus. It was observed that Ce administered at the same time or before E2, was able to inhibit the induction by E2 of fetal thymidine kinase (TK-F), of creatine kinase of brain-type (CK-BB) and of progesterone receptor (PR). When Ce was given after E2, its inhibitory effects were less evident. Results seemed to indicate that Ce could bind the acceptor sites for E2 receptor and thus inhibit the activity of E2-regulated genes. This assumption was corroborated by the fact that Ce did not modify the activity of enzymes not submitted to E2 regulation.

The thermal stability at 37 degrees C of several clinically relevant enzymes and isoenzymes was assessed by measuring changes in enzyme activity as a function of time under incubation and reaction conditions. Selwyn plots were used in the reaction-condition assessments. Except for CK-1 (BB), all the enzymes investigated are stable enough at 37 degrees C to permit assay. These enzymes were LDH-1, LDH-5, s-AspAT, m-AspAT, apo-s-AspAT, apo-m-AspAT, ALP-liver, ALP-bone, ALP-intestine, ALT, apo-ALT, CK-2, and CK-3. CK-1 is stable at 37 degrees C under assay conditions but not under incubation conditions. We specifically avoided using Arrhenius plots to evaluate thermal stability and point out pitfalls inherent in their indiscriminate use.

NMR-spectroscopy (31P-NMR, 23Na-NMR) has been used to characterize the energy metabolism of cultured endothelial cells of the pig aorta and to determine the energy requirements of the Na-K-ATPase relative to total cellular energy consumption. Endothelial cells exhibited high concentrations of creatine phosphate and cardiac microvascular creatine kinase (CK) was of the BB-CK subtype. BB-CK can therefore be used as a marker-enzyme of endothelial cells in muscular organs. The transmembrane flux-rate of sodium was 117 mumol Nalmin/mg prot in the steady state at an intracellular sodium concentration of 25 mmol/l. Combining microcalorimetric measurements with 23Na-NMR data revealed that the energy requirement of endothelial Na-K-ATPase despite the high surface to volume ratio comprises only 3-5% of total cellular energy consumption.

An increase in serum creatine kinase-MB (CK-MB) isoenzyme is regarded as a specific indicator of acute myocardial infarction. We analyzed retrospectively the clinical data of 94 patients whose serum creatine kinase MB (CK-MB) was measured with immunoinhibition kit which measures all residual CK activity following inactivation of M-subunit. There were 21 patients with chronic obstructive lung disease (COLD), 17 patients with pneumonia, 17 patients with pulmonary tuberculosis (TBC), 16 patients with non-small cell lung cancer (NSCLC), 10 patients with small cell lung cancer (SCLC), 8 patients with malignancies of other origin (NPL), and 5 patients with chronic heart diseases. The results revealed that serum concentrations of CK-MB in SCLC, NSCLC and TBC were significantly greater than those in other groups (P < 0.1). Clinical examination showed no evidence of myocardial infarction, injury, or tumor involvement of the heart. We assumed that those results are due to interference of the CK-BB isoenzyme in the immunoinhibition method. So we suggest that in clinical practice, markedly elevated levels of CK-MB measured with immunoinhibition kit, after the exclusion of the myocardial injury, may point toward the existence of a malignancy or TBC of the lungs.

Cardiac markers are used to evaluate functions of heart. However, there are no satisfactory cardiac biomarkers for the diagnosis of acute myocardial infarction (AMI) within 4 h of onset of chest pain. Among novel cardiac markers, glycogen phosphorylase BB (GPBB) is of particular interest as it is increased in the early hours after AMI. The present study was conducted with the objective to find out the sensitivity and specificity of GPBB over other cardiac markers i.e. myoglobin and CKMB in patients of AMI within 4 h after the onset of chest pain. The study includes 100 AMI patients and 100 normal healthy individuals as controls. In all the cases and controls, serum GPBB and myoglobin concentrations were measured by ELISA where as CK-MB was measured by diagnostic kit supplied by ERBA. The sensitivity and specificity of glycogen phosphorylase BB (GPBB) were greater than CK-MB and myoglobin in patients of AMI within 4 h after the onset of chest pain. Hence, glycogen phosphorylase BB (GPBB) can be used as additional biomarker for the early diagnosis of AMI.

BACKGROUND

Diagnosis of acute coronary syndrome (ACS) is frequently a challenging task while immediate risk stratification remains crucial for the prompt implementation of appropriate therapy in this setting. The prolonged release pattern of both CK-MB mass and cardiac troponins makes it difficult to identify the origin of recent chest pain, thus a combination of early and later biomarkers might further facilitate the differential diagnosis. The study was designed to evaluate the efficacy of multi-marker approach using biochip array technology in identifying ACS shortly after the symptom onset.

METHODS

The study group consisted of 42 patients suspected for ACS. Subjects were diagnosed as presenting with unstable angina (UA), non-ST-elevation myocardial infarction (NSTEMI) or ST-elevation myocardial infarction (STEMI). Biomarkers in the serum were determined twice: on admission (≤6 hours from the chest pain onset) and after next 6 hours. Cardiac troponin I was measured by routine sensitive automated assay (STAT cTnI) while other 6 cardiac markers (heart-fatty acid binding protein - H-FABP, myoglobin, glycogen phosphorylase BB, cTn I, CK-MB mass and carbonic anhydrase III) were assessed using biochip array technology.

RESULTS

STAT cTnI concentrations within 6 hours from the symptom onset were elevated over the 99(th) percentile for reference population in 83.3% of subjects but none reached the cut-off value for myocardial infarction. Instead, H-FABP demonstrated a very good efficacy in early detection of ACS (90.5%), better than myoglobin and CK-MB mass. Sensitivity of H-FABP calculated for NSTEMI/STEMI subjects reached 100%. The diagnostic efficacy of troponin, myoglobin and CK-MB mass assay markedly increased within 12 hours. It was only for the patients with UA that the cardiac panel was not efficient in the early stratification of risk.

CONCLUSIONS

A multi-marker strategy with H-FABP and highly sensitive troponin included enhances the early diagnosis and decision making process in patients with ACS. A new biochip cardiac array technology may serve as a powerful tool for ACS detection in the clinical practice.

OBJECTIVE

To investigate the changes and the clinical significance of N-terminal pro-brain natriuretic peptide (NT-proBNP) and glycogen phosphorylase isoenzyme BB (GPBB) levels in neonates with asphyxia complicated by myocardial injury.

METHODS

Sixty-four neonates with asphyxia (39 mild, 25 severe) were enrolled. Of the 64 neonates, 30 had myocardial injury and 34 did not develop myocardial injury. Twenty-five healthy neonates served as a control group. Plasma levels of NT-proBNP and GPBB were measured using ELISA. Myocardial enzymes and cardiac troponin I were stimultaneously measured, and electrocardiography and chest radiographs were obtained.

RESULTS

The plasma levels of NT-proBNP and GPBB in neonates with myocardial injury were significantly higher than those in neonates without myocardial injury and in the control group (P<0.01). The neonates with severe asphyxia had significantly increased plasma NT-proBNP and GPBB concentrations compared to those with mild asphyxia and the control group (P<0.01). Spearman rank correlation analysis showed that plasma NT-proBNP level was positively correlated with plasma GPBB level in neonates with asphyxia. Plasma levels of NT-proBNP and GPBB were also positively correlated with plasma levels of CK-MB, CK and LDH (P<0.01).

CONCLUSIONS

Both NT-proBNP and GPBB can be used as biomarkers of myocardial injury in neonates with asphyxia. The measurement of plasma NT-proBNP and GPBB levels was useful in early identification of myocardial injury and severity evaluation in neonates with asphyxia.

Possibilities of recognizing cerebral function disturbances in newborns as early as the acute phase of the condition, and of prognosticating permanent brain damage are described. Clinical findings such as lethargy or spasms in the second and third weeks of life following hypoxia indicate severe function disturbances with poor prognosis. In contrast, the same symptoms in the first week after a hypoxia or following a cerebral hemorrhage are only of relatively slight prognostic significance. Metabolic parameters such as lactate, hypoxanthine, CK-BB, and ammoniemia are elevated in the first few hours and days after a perinatal hypoxia and correlate with the current disturbances in the central nervous system. However, they are not indicative of a possible occurrence of later brain damage. Cerebral sonography furnishes excellent information concerning gross structural changes and cerebral hemorrhages. The extent of the hemorrhages, classified in 4 stages, correlates well with the later development. Less severe functional disturbances can also be identified by electroencephalography together with observation of behavior (polygraphy). Here also, findings correlate well with the children's subsequent development. The combination of sonographic findings and neurophysiological diagnosis is at present the best source of information concerning the later occurrence of brain damage.

Silver nanoparticles (AgNPs) are widely applied in various aspects of life. However, recent studies reported their potential toxicity both on environment and human health. The present study aimed to unravel the underlying molecular mechanisms involved in AgNPs-induced brain toxicity. Moreover, chemopreventive effect of tranilast, an analogue of tryptophan metabolite and a mast cell membrane stabilizer was evaluated. Thirty Sprague Dawley rats were enrolled equally into Normal control group, AgNPs-intoxicated group (50 mg/kg, 3 times/week) and tranilast (300 mg/kg, 3 times/week)+AgNPs group. AgNPs administration triggered brain oxidative stress as depicted by reduced Nrf-2 expression, decreased TAC and GSH as well as upregulated brain lipid peroxidation. The apparent brain oxidative damage was accompanied by elevated levels of inflammatory cytokines (IL-1β, IL-6 and TNF-α). Moreover, brain levels of TLR4, NLRP3 and caspase-1 were up-regulated. Additionally, histological study indicated marked cellular injury in cerebrum and cerebellum specimens. This was concomitant with elevated serum CK activity and CK-BB level. On the other hand, tanilast administration remarkably alleviated AgNPs-induced brain toxicity. The present study shed the light on implication of TLR4/NLRP3 axis and NrF2 in AgNPs-induced brain toxicity. In addition, it explored the potential protective effect of tranilast on AgNPs-induced brain injury via antioxidant and anti-inflammatory efficacies.

Keywords: NLRP3; Nrf-2; Silver nanoparticles; Tranilast; cerebral toxicity.

In this study, we investigated the effectiveness and microbial mechanism of Bacillus amyloliquefaciens biofertilizer on reducing ammonia volatilization in farmland soil. Pot experiments were carried out to explore the effects of B. amyloliquefaciens biofertilizer (BB) and chemical fertilizer on soil ammonia volatilization, crop yield and quality, and soil microbial community. Four fertilization strategies were tested, namely no fertilizer (CK), 100% chemical fertilizer (C), 50% BB and 50% chemical fertilizer (B1), and 100% BB (B2). The dynamic flow-through chamber method was used to determine the soil ammonia volatilization flux after fertilization. The soil bacterial community during the peak period of ammonia volatilization was analyzed using 16S rDNA high-throughput sequencing. The results showed that the amount of ammonia volatilization in B1 and B2 decreased by 79.5% and 84.8%, respectively, as compared with treatment C. B2 had the lowest nitrate content and the highest yield; the yield of B2 increased by 50.5% and 12.3% as compared to that of CK and C, respectively. B1 had the highest content of vitamin C, which was 67.6 mg ·kg^-1. The application of BB improved the diversity and richness of soil bacterial community, especially the relative abundance of Bacillus and Nitrospira. This shows that BB plays an important role in preventing air pollution and improving nitrogen utilization.

Keywords: Bacillus amyloliquefaciens; ammonia volatilization; biofertilizer; plant growth promoting rhizobacteria; soil bacterial community.

Using malachite green single agent coloration and acetate membrane electrophoresis, we studied the cellular creatine kinase (CK) activity and its isoenzymes in 7 normal controls and 26 leukemia patients. The leukemic cellular CK activity was 12.62 +/- 4.86 u/mg protein, 2.2 times higher than the normal value (5.73 +/- 2.66 u/mg protein, p less than 0.05). Only 2 of 5 normal leukocyte samples showed '+' CK isoenzyme MM. 22 leukemia patients had CK isoenzyme. CK-BB appeared mainly in acute granulocytic leukemic, and CK-MM mainly in other types. CK-MB was also found in 6 patients. The recurrence of CK-BB may indicate atavism, and the enhanced anaerobic glycolysis and the accelerated energetic turnover may be on of the metabolic characteristics of leukemic cell.

Rhabdomyosarcomas have been classified on the basis of their degree of differentiation, a feature closely related with their response to chemotherapy. We recently reported the role of creatine kinase isoenzymes as tumoral markers in the diagnosis of rhabdomyosarcoma. The present study was designed to determine whether these isoenzymes are also good markers of the degree of differentiation of these neoplasm. Dimethyl sulfoxide, a well-known differentiating agent, was used to induce myogenic differentiation. This agent increased CK-MM and/or CK-MB fractions, and decreased CK-BB isoenzyme in rhabdomyosarcoma cell lines. The variable behavior of MCK-1 macromolecule was unrelated to the degree of differentiation. Our findings suggest that CK-MM, CK-MB and CK-BB isoenzymes are good markers of the degree of differentiation in rhabdomyosarcomas and could be used as specific markers of prognostic and diagnostic value.

OBJECTIVE

Total phosphoglycerate mutase (PGM) activity in serum has been shown to be increased in acute myocardial infarction with the same time course as creatine kinase (CK) activity. However, the increase in the muscle (MM) and in the cardiac (MB) PGM isoenzymes was not as high as expected. The present study was undertaken to characterise PGM inactivation by serum and to compare it with serum CK inactivation.

METHODS

The PGM and the CK activities of extracts of human heart, skeletal muscle, and brain were determined spectrophotometrically after incubation with different media, namely: plasma, whole serum, dialysed serum, heated serum, serum ultrafiltrate, urate solution, and buffer solution.

RESULTS

Type MM PGM was inactivated by plasma, whole serum, heated serum, dialysed serum, and serum ultrafiltrate. Inactivation in dialysed serum was reduced by EDTA and largely reversed by thiol agents. Inactivation in serum ultrafiltrate was not prevented by EDTA and only partially reversed by dithiothreitol. The muscle and type BB CK isoenzymes were inactivated in all the tested media. The incubation of human and rabbit skeletal muscle PGM and CK in urate solution showed that urate does not affect mutase activity under conditions that inactivate CK.

CONCLUSIONS

These results confirm the mechanisms of CK inactivation proposed by others and show that the type M PGM subunit is inactivated by two different mechanisms, which appear to involve the thiol groups of the enzyme. One mechanism is caused by either a protein component or a protein bound serum component and involves calcium ions and/or another chelatable metal ion. The other mechanism is caused by a lower molecular weight serum component and is metal ion independent.

We investigated the effect of nuclear factor of activated T cells c1 (NFATc1) on the growth and vascular generation of human ovarian carcinoma SKOV3 cell-transplanted tumors in nude mice and explored the possible underlying mechanism. NFATc1 siRNA was transfected into the SKOV3 cells, which were then subjected to immunofluorescence tests and real-time reverse transcription polymerase chain reaction (RT-PCR) to determine the transfection-induced inhibition rate. The tumor volumes in the nude mice in all groups were measured to determine the in vivo antitumor effect of NFATc1 siRNA. Immunohistochemical (IHC) methods were employed to detect NFATc1 expression in tumor tissue, combined with cytokeratin (CK) staining to label the epithelial origin of the tumor tissue. CD34 and podoplanin were used as markers for labeling microvessels and microlymphatic vessels, respectively. The densities of microvessels and microlymphatic vessels in each group were calculated and statistically analyzed. RT-PCR and western blotting were performed to detect the protein and mRNA expression levels of NFATc1, the ELR+ CXC chemokine interleukin (IL)-8, fibroblast growth factor-2 (FGF-2), and platelet-derived growth factor BB (PDGF BB) in xenografted tumor tissue in all groups. NFATc1 was highly expressed in tumor tissue in the control groups. The intervention group exhibited a tumor growth inhibition rate of 57.08% and presented a lower tumor weight and volume compared with the two control groups. In the control groups, the microvessel densities were 12.00 ± 1.65 and 11.47 ± 0.32, respectively, and the microlymphatic vessel densities were 10.03 ± 0.96 and 9.95 ± 1.12; these values were significantly higher than in the intervention group. RT-PCR and western blot shows that NFATc1 siRNA could markedly suppress the expression of IL-8, FGF-2 and PDGF BB at the mRNA and the protein level. In conclusion, it was shown that NFATc1 siRNA significantly suppresses the growth and vascular generation of SKOV3 human ovarian carcinoma cell-transplanted tumors subcutaneously xenografted into nude mice. The downregulation of the expression of IL-8, FGF-2 and PDGF BB may be one of the mechanisms underlying the above inhibitory effects.

Objective: To explore the clinical related factors of neonatal hand-foot-mouth disease (HFMD) complicated with encephalitis. Method: The neonatal HFMD complicated with encephalitis treated in our hospital from July 2015 to July 2020 was taken as the object of study. According to the NBNA score at discharge, the patients were divided into normal group and abnormal group. The clinical symptoms, auxiliary examination and prognosis of the two groups were compared. Result: (1) General condition: there was no significant difference in sex, age, duration of fever, treatment time and etiological test between the two groups (P > 0.05). (2) Clinical symptoms and signs: there was significant difference in abnormal consciousness between the two groups (P < 0.05). However, there was no significant difference in skin rash, respiratory system symptoms, digestive system symptoms, signs of high intracranial pressure, increased muscle tone and weakening of primitive reflex (P > 0.05). (3) Auxiliary examination: the number of white blood cells and the level of cytokines (CK-BB, UCH-L1) in cerebrospinal fluid (CSF) in the group with abnormal NBNA score were significantly higher than those in the group with normal NBNA score (P < 0.05). The serum IgM level in the abnormal NBNA score group was higher than that in the normal NBNA score group, and the serum IgG level in the abnormal NBNA score group was lower than that in the normal NBNA score group, and the difference was statistically significant (P < 0.05). The abnormal rate of Craniocerebral MRI in abnormal NBNA score group was higher than that in normal NBNA score group, and there was significant difference between the two groups (P < 0.05). There was no significant difference in the levels of protein, sugar, chloride, lactate dehydrogenase, and MMP-9 in CSF and the abnormal rate of amplitude integrated EEG (aEEG) between the two groups (P > 0.05). (4) The prognoses of patients with normal and abnormal NBNA score are good, and there are not significantly differences in the prognosis between the two groups (P > 0.05). Conclusion: (1) Neonatal HFMD complicated with encephalitis occurs more than 10 days after birth, there is no obvious abnormality in male and female, the vast majority of newborns have febrile symptoms, rash is not its specific manifestation, and most of them are atypical. (2) The positive rate of HFMD-related virus detected in CSF of neonatal HFMD is high. For newborns with abnormal consciousness, CSF examination should be accomplished in time, which has certain clinical significance for early diagnosis and treatment of severe newborns. (3) The increase of white blood cell count and cytokines (CK-BB, UCH-L1) in CSF of neonatal HFMD complicated with encephalitis has a certain clinical reference value for early diagnosis and identification of severe newborns. (4) There is a certain humoral immune disorder in newborns with HFMD complicated with encephalitis, but the overall prognosis is better due to the protective effect of maternal IgG.

Keywords: CSF examination; clinical symptoms; inflammatory factors; neonates HFMD; prognosis; risk factors.