Proximal tubular FHL2, a novel downstream target of hypoxia inducible factor 1, is a protector against ischemic acute kidney injury.

Four-and-a-half LIM domains protein 2 (FHL2) is an adaptor protein that may interact with hypoxia inducible factor 1α (HIF-1α) or ß-catenin, two pivotal protective signaling in acute kidney injury (AKI). However, little is known about the regulation and function of FHL2 during AKI. We found that FHL2 was induced in renal tubular cells in patients with acute tubular necrosis and mice model of ischemia-reperfusion injury (IRI). In cultured renal proximal tubular cells (PTCs), hypoxia induced FHL2 expression and promoted the binding of HIF-1 to FHL2 promoter. Compared with control littermates, mice with PTC-specific deletion of FHL2 gene displayed worse renal function, more severe morphologic lesion, more tubular cell death and less cell proliferation, accompanying by downregulation of AQP1 and Na, K-ATPase after IRI. Consistently, loss of FHL2 in PTCs restricted activation of HIF-1 and ß-catenin signaling simultaneously, leading to attenuation of glycolysis, upregulation of apoptosis-related proteins and downregulation of proliferation-related proteins during IRI. In vitro, knockdown of FHL2 suppressed hypoxia-induced activation of HIF-1α and ß-catenin signaling pathways. Overexpression of FHL2 induced physical interactions between FHL2 and HIF-1α, ß-catenin, GSK-3ß or p300, and the combination of these interactions favored the stabilization and nuclear translocation of HIF-1α and ß-catenin, enhancing their mediated gene transcription. Collectively, these findings identify FHL2 as a direct downstream target gene of HIF-1 signaling and demonstrate that FHL2 could play a critical role in protecting against ischemic AKI by promoting the activation of HIF-1 and ß-catenin signaling through the interactions with its multiple protein partners.

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry: An effective method for identification and phylogenetic analysis of Leishmania species.

BACKGROUND: Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a rapid and reproducible method that has been widely applied for the identification of bacteria and fungi. However, this technique has not yet been applied in clinical laboratories for parasitology, such as for the study of the protozoan Leishmania. METHODOLOGY: By using MALDI-TOF MS, mass spectra database entries (MSPs) were created with 7 World Health Organization reference strains in order to establish a rapid method for Leishmania species identification. Furthermore, cluster analysis was performed with 18 Chinese Leishmania isolates. PRINCIPAL FINDINGS: The MSPs of Leishmania corresponded well with our past identification results, and the dendrogram analysis result was more or less similar to that of the phylogenetic analysis performed by multi-locus sequence typing. CONCLUSIONS/SIGNIFICANCE: MALDI-TOF MS is a promising method that offers both rapidity and efficiency for the identification and dendrogram analysis of Leishmania species.

Performance of a new Candida anti-mannan IgM and IgG assays in the diagnosis of candidemia.

Candida is one of the most frequent pathogens of bloodstream infections, which is associated with high morbidity and mortality rates. Rapid immunological detection methods are essential in the early diagnosis of candidemia. Anti-mannan is one of host-derived biomarkers against cell wall components of Candida. We conducted this study to evaluate the diagnostic performance of two anti-mannan assays (IgM, IgG) for candidemia through the analysis of 40 candidemia patients, 48 participants with Candida colonization and 213 participants with neither Candida colonization nor Candida infections (13 patients with other bloodstream infections, 145 hospitalized patients and 55 healthy controls). The performance of the two assays were evaluated by calculating their sensitivity and specificity. The sensitivity ranged from 0.78 to 0.80 for the IgM assay and 0.68 to 0.75 for the IgG assay. The specificity ranged from 0.97 to 0.98 for the IgM assay and 0.91 to 0.94 for the IgG assay. The diagnostic performance of the anti-mannan IgM assay was better than that of IgG, with higher sensitivity and specificity. Combining the two assays (positive results of single or both assays are both considered as positive) could improve the sensitivity up to 0.93 (0.79-0.98) and only slightly reduce the specificity (0.93(0.89-0.95)). The anti-mannan IgM, IgG assays are rapid and cost-effective assays that may be probably useful in the diagnosis of candidemia.

[Preliminary Study on PCR Method for Laboratory Diagnosis with Fecal Specimens of Entamoeba histolytica Infection].

OBJECTIVE: To develop a PCR method for Entamoeba histolytica（ E.histolytica) detection in fecal specimens, and to compare the performance of PCR to that of microscopy and ELISA. METHODS: Two pairs of self-designed primers and 2 pairs of primers from references based on small subunit ribosome RNA (SSU rRNA) fragment of E. histolytica standard strain were synthetized. DNA from E. histolytica reference strain were amilified by the conventional PCR using the 4 pairs of primers. 221 stool samples from diarrhea patients were collected and detected for E. histolytica by three methods: Entamoeba trophozoites and cysts detection by microscopy, E. histolytica-specific antigen detection using enzyme-linked immunosorbent assay (ELISA) kit ( E. HISTOLYTICA II), amplification of SSU rRNA fragment of E. histolytica by PCR method. Positive rate of three methods were compared by chi-square test, and Kappa test was applied to determine the concordance among the three methods. RESULTS: Specific fragments of E. histolytica were amplified by the PCR method we developed in this study. Positive rates of PCR, microscopy and ELISA were 2.26%, 0.90% and 9.50%, respectively. The positive rates of the three methods were significantly different ( χ 2 =23.34, P<0.01). The Kappa value of PCR and microscopy was 0.216, and that of PCR and ELISA method was -0.134, both of which showed a weak consistency. PCR results showed best consistency with clinical diagnosis. CONCLUSION: The PCR method we established in this study has a better performance in accuracy than microscopy and ELISA have in laboratory diagnosis of E. histolytica infection.

Case Report: Morphologic and Genetic Identification of Cerebral Sparganosis.

A 50-year-old Chinese woman with a history of weakness and paroxysmal seizures of the left limb presented to our hospital with a ten-day history of neck pain. Imaging showed that there was a mass in the frontal lobe of her brain. On resection of the mass, a motile worm was identified. Morphological observation and molecular analysis of the mitochondrial COX1 and 28S rRNA genes of the worm extracted from the brain identified the causative agent as Spirometra mansoni. Homology search of the polymerase chain reaction (PCR)-amplified products from the case was conducted against gene fragments from local wild frogs. High homology was found between them, showing her likely exposure was frog consumption.

A rapid high-resolution melting method for differentiation of Leishmania species targeting lack gene.

OBJECTIVES: The aim of this research is to verify that if lack gene can be used for differentiation of Leishmania under HRM assay. METHODS: Two specific primers were designed targeting polymorphic sites on the lack gene sequence. DNA from promastigotes of six species of Leishmania based on reference strains were tested following a HRM protocol. We also tested ten Chinese isolates in blind to validate our method. RESULTS: Combined with amplicon of the two primers, the six reference strains can be easily discriminated without the effect of initial concentration of DNA templates. Ten Chinese isolates detected by our HRM method resulted in full accord with the standard identification results in previous study. CONCLUSION: HRM is a rapid and reproducible method to discriminate different Leishmania species and lack gene is a potential novel biological characteristic for easy differentiation of Leishmania isolates in China.