[Clinical features of children with Cunninghamella spp. infection: a case report and literature review].

We report a case of mucormycosis induced by Cunninghamella spp. infection in a ten-year-old girl with acute lymphoblastic leukemia, who developed fever and respiratory symptoms after chemotherapy and was diagnosed with invasive fungal disease. Peripheral blood DNA sequences were analyzed using metagenomic next-generation sequencing (mNGS), and by comparison with the Pathogens Metagenomics Database (PMDB), we identified Cunninghamella spp. with sequence number 514 as the pathogen. The patient was treated with amphotericin B combined with posaconazole and showed a favorable response. We searched Pubmed, Embase, CNKI, and Wanfang database for reports of cases of Cunninghamella spp. infection in children and retrieved 22 reported cases (including 12 males) with a median age of 13.5 (3-18) years. In these 22 cases, hematological malignancy was the most common underlying condition (19/22), and most of patients experienced an acute onset and rapid progression with respiratory symptoms (14/20) and fever (16/20) as the most common symptoms. CT imaging often showed unilateral lesions with varying imaging findings, including pulmonary nodules or masses, infiltrative changes, and pleural effusion. Definite diagnoses were established in 18 of the cases, and 4 had probable diagnoses; the lungs and skin were the most frequent organs compromised by the infection. A definite diagnosis of Cunninghamella spp. infection still relied on histopathological examination and fungal culture, but the molecular techniques including PCR and mNGS had shown potentials in the diagnosis. Almost all the cases received antifungal treatment after diagnosis (21/22), and 13 patients also underwent surgeries. Death occurred in 9 (42%) of the cases at a median of 19 (4-54) days after onset of the signs or symptoms. The patients receiving antifungal therapy combined with surgery had a high survival rate (9/13, 69%) than those with antifungal therapy alone (3/8, 37%). Invasive fungal disease is a common complication in immunoco-mpromised patients, but Cunninghamella spp. infection is rare and has a high mortality rate. In cases highly suspected of this disease, active diagnosis and early treatment are critical to improve the survival outcomes of the patients.

[Clinical analysis of lymphocytic thyroiditis coexistent with papillary thyroid carcinoma].

OBJECTIVE: To analyse the clinical features and treatment strategies of papillary thyroid carcinoma(PTC) coexistent with lymphocytic thyroiditis (LT). METHODS: A total of 292 patients including 25 males and 267 females with LT and thyroid nodules treated in the department of head and neck surgery between Sep 2011 and Sep 2014 was analysed retrospectively and divided into two groups according to pathological results, of them 262 patients, with a median age of 47 years old, were LT with PTC and 30 patients, with a median age of 54 years old, were LT with benign nodules. Among 262 patients having LT with PTC, 259 were diagnosed as having malignant or suspicious malignant nodules and 3 having benign nodules with ultrasound before surgery, 98 cases were considered multifocal malignant nodules by preoperative ultrasound, and 112 cases were positive in cervical lymph nodes, including bilateral positive in 37 cases. Of 30 patients having LT with benign nodules, 14 were diagnosed malignant or suspicious malignant nodules and 16 benign nodules. RESULTS: The mean age in 262 patients with PTC was less significantly than that in 30 patients with benign nodules (P<0.05). Ultrasound showed a high proportion of calcification and microcalcification in patients with PTC (34%) compared to patients with benign nodules(13%)(P<0.05). There were not significant differences in the mean levels of serum thyroid stimulating hormone(TSH) (2.80 vs 2.99 mU/L, P=0.233), thyroglobulin(TG) (27.14 vs 18.60 µg/L, P=0.747), and anti-thyroglobulin antibodies(ATG)(417.3 vs 378.7 U/ml, P=0.834) between patients with PTC and those with benign nodules. In patients with PTC, multifocal tumor accounted for 42%. The central and lateral lymph node metastasis rates were respectively 50% and 24%, and the occult metastasis rate of lateral neck lymph node was 16%. Univariate analysis showed that age less than 45 years old, multifocal tumor, tumor diameter more than or equal to 2cm and extrathyroidal extension were associated with central lymph node metastasis (P<0.05), but not with lateral neck metastasis. Multivariate analysis showed a closed correlation only between the lymph node metastases in central and lateral neck levels (P<0.05). CONCLUSIONS: Calcification and microcalcification have the same importance in the ultrasonic diagnosis for PTC in patients with LT. Total thyroidectomy and prophylactic central lymph node should be a choice for LT with PTC. Lymph node metastasis in level Ⅵ indicates the possibility of lateral cervical lymph node metastasis in the patients having LT with PTC.

A novel marine drug, SZ-685C, induces apoptosis of MMQ pituitary tumor cells by downregulating miR-200c.

OBJECTIVE: We found a novel marine drug, SZ-685C, that was isolated from the secondary metabolites of a mangrove endophytic fungus (No. 1403) collected from the South China Sea, which has been reported to inhibit the proliferation of certain tumor cells. However, its anticancer mechanism remains unknown. The aims of this study were to observe the effectiveness of SZ-685C on pituitary adenoma cells and determine the underlying mechanisms of action. METHODS: A rat prolactinoma cell line, MMQ, was used in this study. A dose escalation of SZ-685C was performed on this cell line, and cell viability was assessed using an MTT assay. Hoechst 33342, Annexin V-FITC/PI, TUNEL staining and flow cytometry were used to evaluate the extent of apoptosis at each concentration of SZ-685C. The effect of SZ-685C on prolactin expression was also evaluated using RT-PCR and immunoblotting. Quantitative RT-PCR was used to detect the expression of miR-200c in SZ-685C-stimulated MMQ cells and pituitary adenoma tissues. This miRNA was then overexpressed in MMQ cells via transfection of a miR-200c mimic to identify the mechanism underling the anti-tumor effect of SZ-685C. RESULTS: SZ-685C inhibited MMQ cell growth in a dose-dependent manner but showed little toxicity toward rat pituitary cells (RPCs). The IC50s of SZ-685C in MMQ cells and RPCs were 13.2 ± 1.3 mM and 49.1 ± 11.5 mM, respectively, which was statistically significant. Increasing numbers of apoptotic cells were observed in response to escalating concentrations of SZ-685C, and the expression level of prolactin (PRL) was inhibited. Nevertheless, the level of PRL mRNA was unchanged. Additionally, miR-200c was upregulated in MMQ cells compared with RPCs, and downregulation of miR- 200c was observed in SZ-685C-treated MMQ cells. Furthermore, the overexpression of miR-200c weakened the effect of SZ-685C-induced apoptosis of MMQ cells. CONCLUSIONS: Our results suggest that SZ-685C induces MMQ cell apoptosis in a miR-200c-dependent manner. Therefore, SZ-685C might be a useful alternative treatment for pituitary adenoma.

A new nonadride derivative from mangrove fungus (strain No. k38).

A new nonadride derivative, ( - )-1-hydroxybyssochlamic acid (1) and the known ( - )-byssochlamic acid (2) were isolated from mangrove fungus (strain No. k38) collected from the South China Sea coast. The structure and relative configuration of 1 were elucidated by spectral data and X-ray diffraction analysis. Primary bioassays showed that 2 had medium cytotoxic activity against HEp-2 and HepG2 Cells, and 1 exhibited weak activity.

A mathematical model for synergistic eukaryotic gene activation.

The precise biochemical mechanism underlying the synergistic action of gene activators on eukaryotic transcription has eluded a solution, largely because of the technical difficulties inherent in analyzing the mechanics of a 2.5 MDa complex comprising greater than 50 polypeptide components. To complement the biochemical approach we have employed mathematical modeling as a means to understand the mechanism of synergy. Parameters relevant to activated transcription were varied in a simple biochemical system and the data were compared to the transcriptional response predicted by a multi-component statistical model. We found that the model achieved a consistent, semi-quantitative description of the measured transcriptional response, and enabled the characterization and measurement of thermodynamic parameters in the in vitro system. The results provide evidence for the existence of cooperativity in the activation process beyond what would be predicted from one current model suggesting that activators function solely by simple recruitment of the general transcription machinery to the promoter.

Aminobenzoic acid compounds as HOCl traps for activated neutrophils.

This study was designed to develop traps for hypochlorous acid (HOCl) which could be used to detect HOCl in the microenvironment of activated neutrophils. Reagent HOCl was found to react with para-aminobenzoic acid (PABA) in aqueous solution to produce a predominant metabolite detectable by high performance liquid chromatography (HPLC). Mass spectroscopy and nuclear magnetic resonance identified this metabolite as the ring addition product 3-chloro PABA. The related compound para-aminosalicylic acid (PAS) was also metabolized by HOCl to 3-chloro PAS. The formation of the 3-chloro metabolite was specific for reactions involving HOCl, since several other oxidants in chloride buffer failed to produce the metabolite. Human blood neutrophils activated by phorbol myristate acetate or zymosan in the presence of PABA (or PAS) used their HOCl to produce large amounts of the 3-chloro metabolite. The formation of 3-chloro PABA was inhibited by azide, catalase, and taurine, which is consistent with the production of the metabolite by the neutrophil myeloperoxidase (MPO) pathway. The reaction of HOCl with PABA and PAS was relatively slow as shown by competitive reactions with endogenous antioxidants like taurine, methionine, and glutathione. This was confirmed in reactions involving PABA/PAS and reagent HOCl or HOCl generated by the MPO enzyme system. In these in vitro systems, glutathione and serum completely inhibited the formation of the 3-chloro metabolite. In contrast, activated neutrophils metabolized PABA/PAS to the 3-chloro metabolite even in the presence of 1% serum. These findings demonstrate that PABA and PAS are specific trapping agents for HOCl produced by neutrophils in complex biological conditions.

Tumor necrosis factor increases the elastolytic potential of adherent neutrophils: a role for hypochlorous acid.

Neutrophils adhered to biologic surfaces exhibit proteolytic cleavage of surface proteins even in the presence of proteinase inhibitors. Such proteolysis is restricted to the pericellular space and appears to require the dual action of proteinases and reactive oxygen species. The present study was designed to investigate the mechanism by which tumor necrosis factor-alpha (TNF) stimulates neutrophil proteolysis. Tissue culture wells were coated with insoluble 3H-labeled elastin substrate. Human blood neutrophils (0.5 to 2.0 x 10(6) cells/ml/well) were incubated in the coated wells for 4 to 18 h at 37 degrees C in the presence of varying concentrations of serum or purified alpha 1-antitrypsin (A1AT). TNF (0 to 1,000 U/ml) was also present in the incubations. Elastin degradation was determined as soluble 3H-elastin fragments released into the supernatants. As previously reported, cells (no TNF) exhibited spontaneous elastolysis even in the presence of 1% serum or 4 microM AlAT. Compared with cells incubated alone (no TNF), TNF increased elastolysis 3-fold in the 4-h incubations and 83% in 18-h incubations. TNF also significantly increased proteolysis when neutrophils were concurrently treated with phorbol myristate acetate or N-formylmethionylleucylphenylalanine. Since TNF is known to prime neutrophils for hypochlorous acid (HOCl) release, the present study hypothesized that the enhancement of proteolysis by TNF was related to increased release of HOCl. First, TNF caused a 4-fold increase in HOCl release by neutrophils adhered to elastin surfaces. Second, the effect of methionine on elastolysis by adherent neutrophils was studied.(ABSTRACT TRUNCATED AT 250 WORDS)

Endotoxin priming of monocytes augments Fc gamma receptor cross-linking-induced TNF-alpha and IL-1 beta release.

Cross-linking receptors for the Fc region of immunoglobulin G (IgG) (Fc gamma R) induces tumor necrosis factor-alpha (TNF-alpha) release; however, there is controversy about release of interleukin (IL)-1 beta. The purpose of this study was to investigate the role of endotoxin priming on the ability of monocytes to release these cytokines after Fc gamma R cross-linking. Monocytes were incubated with plated or soluble human IgG or albumin with or without endotoxin priming. Monocytes isolated by Percoll, containing low concentrations of endotoxin, and incubated on plated IgG released 4.5 +/- 1.6 ng/ml TNF-alpha and 1.6 +/- 0.6 ng/ml IL-1 beta. Monocytes isolated by a "clumping" technique released 1.0 +/- 0.4 ng/ml TNF-alpha but no IL-1 beta. Priming with endotoxin, which did not affect Fc gamma R expression, resulted in augmented release of TNF-alpha (4.3 +/- 1.3 vs. 0.1 +/- 0.0 ng/ml, P < 0.05) and IL-1 beta (4.0 +/- 1.0 vs. 0.6 +/- 0.3 ng/ml, P < 0.01) when clumped monocytes were incubated on plated IgG vs. plated albumin.

Tumor necrosis factor infusions in humans prime neutrophils for hypochlorous acid production.

Five cancer patients undergoing intravenous infusions of human recombinant tumor necrosis factor (TNF)alpha were evaluated for the effects these infusions had on the priming of circulating neutrophils for hypochlorous acid (HOCl) production. These patients were also studied for changes in temperature, circulating white blood cell counts, blood pressure, and spontaneous monocyte interleukin 1 beta (IL-1 beta) and TNF production. As predicted by previous in vitro studies, patient neutrophils increased their HOCl production to unopsonized zymosan from a baseline of 29.2 +/- 5.9 nmol I- oxidized/4 x 10(6) cells to a peak of 64.2 +/- 9.8 nmol I- oxidized/4 x 10(6) cells at 4 h after TNF infusion (P less than 0.01). Similar increases were also seen at 4 h with phorbol myristic acetate and opsonized zymosan as the stimuli. The priming effect could be reproduced in neutrophils from a normal individual by incubating them with the 30-min serum samples from the infused patients. The ability of this serum to prime neutrophils was completely blocked by a monoclonal anti-TNF alpha-antibody but not by an anti-IL-1 beta antibody. In addition to the priming of their neutrophils, patients also experienced fever, marked hypotension, and an initial fall, followed by rebound to an elevation, in circulating white blood cell counts. The TNF infusions did not produce detectable circulating IL-1 beta nor did they induce significant production of TNF or IL-1 beta by circulating blood monocytes. These studies confirm the role of TNF in producing the signs of sepsis such as hypotension, fever, and leukopenia followed by leukocytosis.(ABSTRACT TRUNCATED AT 250 WORDS)

Tumor necrosis factor primes neutrophils for hypochlorous acid production.

Tumor necrosis factor (TNF) has a weak direct effect on neutrophil oxidative metabolism and primes neutrophils for oxidant release in response to other stimuli. We examined the effect of recombinant human TNF alpha (rTNF alpha) on production of hypochlorous acid (HOCl) by human neutrophils. TNF alone, even at concentrations of 1,000 U/ml, did not stimulate HOCl production. In contrast, rTNF alpha, in a dose-dependent manner, primed neutrophils for HOCl production in response to the weak agent unopsonized zymosan. rTNF alpha concentrations as low as 10 U/ml resulted in a fivefold increase in HOCl in this system. rTNF alpha-primed cells also exhibited increased phagocytosis. Priming in this model system occurred regardless of whether cells were preincubated with rTNF alpha before addition of zymosan or coincubated with both rTNF alpha and zymosan. rTNF alpha priming for HOCl production could not be washed away and required a lag period of approximately 10 min. rTNF alpha priming was not dependent on extracellular Ca2+ and Mg2+. Preincubation experiments demonstrated that rTNF alpha priming was not inhibited by the microfilament blocker cytochalasin B. Although the mechanism remains unclear, these findings demonstrate that rTNF alpha has an important priming effect on the neutrophil myeloperoxidase pathway.

Hydroxylation of salicylate by activated neutrophils.

Salicylates are metabolized in vivo to hydroxylated compounds, including 2,3-dihydroxybenzoic acid and 2,5-dihydroxybenzoic acid (gentisic acid). The present study hypothesized that activated neutrophils represent one pathway for salicylate hydroxylation. Human neutrophils were incubated in medium containing 10 mM salicylate and stimulated with phorbol myristate acetate (PMA) for 1 hr. The cell-free supernatant fractions were analyzed by HPLC. Neutrophils (1 x 10(6) cells) produced 55 +/- 11 ng of gentisic acid. Neutrophils also produced smaller quantities of 2,3-dihydroxybenzoic acid. Antioxidant inhibitor experiments indicated that superoxide dismutase (SOD), heme protein inhibitors, and glutathione blocked gentisic acid formation, whereas catalase, mannitol, and deferoxamine failed to inhibit. Experiments with the reagent hypochlorous acid (HOCl) and the model myeloperoxidase (MPO) enzyme system did not support a role for the MPO pathway in gentisic acid formation. These findings demonstrate that activated neutrophils can hydroxylate salicylate by an unknown pathway. This pathway may contribute to the increased recovery of hydroxylated salicylates in patients with inflammatory disorders.