Cheng-Shyuan Rau

Post-neurosurgical nosocomial bacterial meningitis in adults: microbiology, clinical features, and outcomes

The clinical data of 62 adult patients who suffered post-neurosurgical nosocomial bacterial meningitis, retrospectively collected over a 16-year period, were studied. Cases were divided into two groups based on the date of presentation, the first period being 1986-1993 and the second 1994-2001. Fever and progressive consciousness disturbance were the most consistent clinical features - signs that may also be attributed to other postoperative neurosurgical problems. The common pathogens included Staphylococcus aureus, coagulase negative Staphylococcus, Pseudomonas aeruginosa, Escherichia coli, and Acinetobacter baumannii. An increase in polymicrobial infections and multi-antibiotic resistance during the second period was identified. In the first half of the study, mortality was 22%, and in the second half 36%. Adult post-neurosurgical nosocomial bacterial meningitis has become an important clinical problem. The choice of appropriate empirical antibiotics is challenging and must be guided by an awareness of the relative frequency of various pathogens and the increasing incidence of resistant strains. Although high mortality rates may, in part, be related to the primary brain pathology, early diagnosis and the timely use of antibiotics based on antimicrobial susceptibility testing are essential for survival.

Profiling muscle-specific microRNA expression after peripheral denervation and reinnervation in a rat model.

MicroRNAs (miRNAs) are a class of highly conserved, non-coding RNAs involved in post-transcriptional gene regulation. The muscle-specific miRNAs, miR-1, miR-133a, and miR-206, are expressed in skeletal muscles and have been shown to contribute to muscle development. To profile their expression after sciatic nerve denervation and reinnervation, the soleus muscles of the rats were analyzed with quantitative real-time PCR at 1 week, 1 month, 2 months, and 4 months after the experiments. In addition, a combined approach using computational prediction by the miRanda website and the Agilent Whole Rat Genome 4 x 44 k oligo microarray experiment was performed to investigate the potential target genes of these three miRNAs in the denervated and reinnervated muscles. The results revealed that with the first downregulation of miR-1 and miR-133a within 1 month in the denervated muscle, the expression of miR-1 and miR-133 increased by approximately 2-fold at 4 months after denervation and reinnervation; on the other hand, the expression of miR-206 was significantly increased to approximately 3-fold 1 month later only following reinnervation but not following denervation, and lasted at least for 4 months. The expression pattern of miR-206 was different from that of miR-1 and miR-133a. Notably, two genes (Hnrpu and Npy) and one gene (Ptprd) were potentially regulated both in the denervated and reinnervated muscle by miR-1 and miR-133a, respectively. There were six potential target genes (Hnrpu, Lsamp, MGC108776, Mef2, Npy, and Ppfibp2) of the upregulated miR-206 in the reinnervated muscle. Among these, three (Hnrpu, Npy, and MGC108776) were potentially regulated by both miR-1 and miR-206. Because the Mef2 transcription factor was reported to promote the transformation of type II fast glycolytic fibers into type I slow oxidative fibers, the upregulation of miR-206 with decreased expression of the Mef2 transcript in the 4 month reinnervated muscle, which presented type II fiber predominance 4 months after nerve microanastomosis, might indicate the role of miR-206 in determining the fiber type after peripheral nerve regeneration.

Whole blood-derived microRNA signatures in mice exposed to lipopolysaccharides

BackgroundLipopolysaccharide (LPS) is recognized as the most potent microbial mediator presaging the threat of invasion of Gram-negative bacteria that implicated in the pathogenesis of sepsis and septic shock. This study was designed to examine the microRNA (miRNA) expression in whole blood from mice injected with intraperitoneal LPS.MethodsC57BL/6 mice received intraperitoneal injections of varying concentrations (range, 10–1000 μg) of LPS from different bacteria, including Escherichia coli, Klebsiella pneumonia, Pseudomonas aeruginosa, Salmonella enterica, and Serratia marcescens and were killed 2, 6, 24, and 72 h after LPS injection. Whole blood samples were obtained and tissues, including lung, brain, liver, and spleen, were harvested for miRNA expression analysis using an miRNA array (Phalanx miRNA OneArray® 1.0). Upregulated expression of miRNA targets in the whole blood of C57BL/6 and Tlr4−/− mice injected with LPS was quantified using real-time RT-PCR and compared with that in the whole blood of C57BL/6 mice injected with lipoteichoic acid (LTA) from Staphylococcus aureus.ResultsFollowing LPS injection, a significant increase of 15 miRNAs was observed in the whole blood. Among them, only 3 miRNAs showed up-regulated expression in the lung, but no miRNAs showed a high expression level in the other examined tissues. Upregulated expression of the miRNA targets (let-7d, miR-15b, miR-16, miR-25, miR-92a, miR-103, miR-107 and miR-451) following LPS injection on real-time RT-PCR was dose- and time-dependent. miRNA induction occurred after 2 h and persisted for at least 6 h. Exposure to LPS from different bacteria did not induce significantly different expression of these miRNA targets. Additionally, significantly lower expression levels of let-7d, miR-25, miR-92a, miR-103, and miR-107 were observed in whole blood of Tlr4−/− mice. In contrast, LTA exposure induced moderate expression of miR-451 but not of the other 7 miRNA targets.ConclusionsWe identified a specific whole blood–derived miRNA signature in mice exposed to LPS, but not to LTA, from different gram-negative bacteria. These whole blood-derived miRNAs are promising as biomarkers for LPS exposure.

Profiling Circulating MicroRNA Expression in Experimental Sepsis Using Cecal Ligation and Puncture

The levels of circulating microRNAs (miRNAs) in mice with experimental sepsis induced by cecal ligation and puncture (CLP) were determined using whole blood samples obtained from C57BL/6 mice at 4, 8, and 24 h after CLP; miRNA expression analysis was performed in these samples using an miRNA array. Microarray analysis revealed upregulation of 10 miRNA targets (miR-16, miR-17, miR-20a, miR-20b, miR-26a, miR-26b, miR-106a, miR-106b, miR-195, and miR-451). The expression of these miRNA targets in the whole blood, serum, and white blood cells (WBCs) of CLP mice was quantified using quantitative real-time PCR; these values were compared to those in sham-operated C57BL/6 mice, and the results indicated that these miRNA targets were significantly up-regulated in the whole blood and serum but not in the WBCs. In addition, the levels of these 10 miRNA targets in the serum of Tlr2−/−, Tlr4−/−, and NF-κB−/− mice at 8 h after CLP did not decrease significantly., which indicated that the transcription of these miRNAs was not directly mediated by the TLR2/NF-κB or TLR4/NF-κB pathway, and pathways induced by exposure to the gram-positive or gram-negative bacteria. Immunoprecipitation with the Argonaute 2 ribonucleoprotein complex revealed significantly increased expression of the 10 miRNA targets in the serum of mice after CLP, and the levels of 6 (miR-16, miR-17, miR-20a, miR-20b, miR-26a, and miR-26b) of these 10 miRNA targets increased significantly in exosomes isolated using ExoQuick precipitation solution. In this study, we identified circulating miRNAs that were up-regulated after CLP and determined the increase in the levels of these miRNAs, and our results suggest that circulating Ago2 complexes and exosomes may be responsible for the stability of miRNAs in the serum.

Entrapment neuropathy results in different microRNA expression patterns from denervation injury in rats

BackgroundTo compare the microRNA (miRNA) expression profiles in neurons and innervated muscles after sciatic nerve entrapment using a non-constrictive silastic tube, subsequent surgical decompression, and denervation injury.MethodsThe experimental L4-L6 spinal segments, dorsal root ganglia (DRGs), and soleus muscles from each experimental group (sham control, denervation, entrapment, and decompression) were analyzed using an Agilent rat miRNA array to detect dysregulated miRNAs. In addition, muscle-specific miRNAs (miR-1, -133a, and -206) and selectively upregulated miRNAs were subsequently quantified using real-time reverse transcription-polymerase chain reaction (real-time RT-PCR).ResultsIn the soleus muscles, 37 of the 47 miRNAs (13.4% of the 350 unique miRNAs tested) that were significantly downregulated after 6 months of entrapment neuropathy were also among the 40 miRNAs (11.4% of the 350 unique miRNAs tested) that were downregulated after 3 months of decompression. No miRNA was upregulated in both groups. In contrast, only 3 miRNAs were upregulated and 3 miRNAs were downregulated in the denervated muscle after 6 months. In the DRGs, 6 miRNAs in the entrapment group (miR-9, miR-320, miR-324-3p, miR-672, miR-466b, and miR-144) and 3 miRNAs in the decompression group (miR-9, miR-320, and miR-324-3p) were significantly downregulated. No miRNA was upregulated in both groups. We detected 1 downregulated miRNA (miR-144) and 1 upregulated miRNA (miR-21) after sciatic nerve denervation. We were able to separate the muscle or DRG samples into denervation or entrapment neuropathy by performing unsupervised hierarchal clustering analysis. Regarding the muscle-specific miRNAs, real-time RT-PCR analysis revealed an ~50% decrease in miR-1 and miR-133a expression levels at 3 and 6 months after entrapment, whereas miR-1 and miR-133a levels were unchanged and were decreased after decompression at 1 and 3 months. In contrast, there were no statistical differences in the expression of miR-206 during nerve entrapment and after decompression. The expression of muscle-specific miRNAs in entrapment neuropathy is different from our previous observations in sciatic nerve denervation injury.ConclusionsThis study revealed the different involvement of miRNAs in neurons and innervated muscles after entrapment neuropathy and denervation injury, and implied that epigenetic regulation is different in these two conditions.

MicroRNA profiling in ischemic injury of the gracilis muscle in rats

BackgroundTo profile the expression of microRNAs (miRNAs) and their potential target genes in the gracilis muscles following ischemic injury in rats by monitoring miRNA and mRNA expression on a genome-wide basis.MethodsFollowing 4 h of ischemia and subsequent reperfusion for 4 h of the gracilis muscles, the specimens were analyzed with an Agilent rat miRNA array to detect the expressed miRNAs in the experimental muscles compared to those from the sham-operated controls. Their expressions were subsequently quantified by real-time reverse transcription polymerase chain reaction (real-time RT-PCR) to determine their expression pattern after different durations of ischemia and reperfusion. In addition, the expression of the mRNA in the muscle specimens after 4 h of ischemia and reperfusion for 1, 3, 7, and 14 d were detected with the Agilent Whole Rat Genome 4 × 44 k oligo microarray. A combined approach using a computational prediction algorithm that included miRanda, PicTar, TargetScanS, MirTarget2, RNAhybrid, and the whole genome microarray experiment was performed by monitoring the mRNA:miRNA association to identify potential target genes.ResultsThree miRNAs (miR-21, miR-200c, and miR-205) of 350 tested rat miRNAs were found to have an increased expression in the miRNA array. Real-time RT-PCR demonstrated that, with 2-fold increase after 4 h of ischemia, a maximum 24-fold increase at 7 d, and a 7.5-fold increase at 14 d after reperfusion, only the miR-21, but not the miR-200c or miR-205 was upregulated throughout the experimental time. In monitoring the target genes of miR-21 in the expression array at 1, 3, 7, 14 d after reperfusion, with persistent expression throughout the experiment, we detected the same 4 persistently downregulated target genes (Nqo1, Pdpn, CXCL3, and Rad23b) with the prediction algorithms miRanda and RNAhybrid, but no target gene was revealed with PicTar, TargetScanS, and MirTarget2.ConclusionsThis study revealed 3 upregulated miRNAs in the gracilis muscle following ischemic injury and identified 4 potential target genes of miR-21 by examining miRNAs and mRNAs expression patterns in a time-course fashion using a combined approach with prediction algorithms and a whole genome expression array experiment.

Computer-assisted pedicle screw placement for thoracolumbar spine fracture with separate spinal reference clamp placement and registration

BACKGROUND The objective of the study was to improve the accuracy of computer-assisted pedicle screw installation in the spine. This study evaluates the accuracy of computer-assisted pedicle screw placement with separate spinal reference clamp placement and registration on each instrumented vertebra for thoracolumbar spine fractures. METHODS Postoperative radiographs and CT scans assessed the accuracy of pedicle screw placement in 21 adult patients on each instrumented vertebra. Screw placements were graded as good if the screws were placed in the central core of the pedicle and the cancellous portion of the body. Screw placements were graded as fair if the screws were placed slightly eccentrically, causing erosion of the pedicular cortex, and with less than a 2-mm perforation of the pedicular cortex. Screw placements were graded as poor if screws were placed eccentrically with a large portion of the screw extending outside the cortical margin of the pedicle and with more than a 2-mm perforation of the pedicular cortex. RESULTS A total of 140 image-guided pedicle screws were placed in 21 patients: 78 in the thoracic and 62 in the lumbar spine. Of the 140 pedicle screw placements, 96.4% (135/140) were categorized as good; 3.6% (5/140), fair; and 0% were poor. All 5 fair placement screws were placed in the thoracic spine without any mobility. CONCLUSION Separate registration increases accuracy of screw placement in thoracolumbar pedicle instrumentation. Separate spinal reference clamp placement in the instrumented vertebra provides real-time virtual imaging that decreases the possibility of downward displacement during manual installation of the screw.

Circulating microRNA signatures in mice exposed to lipoteichoic acid

BackgroundPreviously, we had identified a specific whole blood–derived microRNAs (miRNAs) signature in mice following in vivo injection of lipopolysaccharide (LPS) originated from Gram-negative bacteria. This study was designed to profile the circulating miRNAs expression in mice exposed to lipoteichoic acid (LTA) which is a major component of the wall of Gram-positive bacteria.ResultsC57BL/6 mice received intraperitoneal injections of 100 μg of LTA originated from Bacillus subtilis, Streptococcus faecalis, and Staphylococcus aureus were killed 6 h and the whole blood samples were obtained for miRNA expression analysis using a miRNA array (Phalanx miRNA OneArray® 1.0). Up-regulated expression of miRNA targets in the whole blood, serum and white blood cells (WBCs) of C57BL/6 and Tlr2−/− mice upon LTA treatment in 10, 100, or 1000 ug concentrations was quantified at indicated time (2, 6, 24, and 72 h) using real-time RT-PCR and compared with that in the serum of C57BL/6 mice injected with 100 ug of LPS. A significant increase of 4 miRNAs (miR-451, miR-668, miR-1902, and miR-1904) was observed in the whole blood and the serum in a dose- and time-dependent fashion following LTA injection. Induction of miRNA occurred in the serum after 2 h and persisted for at least 6 h. No increased expression of these 4 miRNAs was found in the WBCs. Higher but not significant expression level of these 4 miRNAs were observed following LTA treatment in the serum of Tlr2−/−against that of C57BL6 mice. In contrast, LPS exposure induced moderate expression of miR-451 but not of the other 3 miRNA targets.ConclusionsWe identified a specific circulating miRNA signature in mice exposed to LTA. That expression profile is different from those of mice exposed to LPS. Those circulating miRNAs induced by LTA or LPS treatment may serve as promising biomarkers for the differentiation between exposures to Gram-positive or Gram-negative bacteria.

Weight-reduction through a low-fat diet causes differential expression of circulating microRNAs in obese C57BL/6 mice

BackgroundTo examine the circulating microRNA (miRNA) expression profile in a mouse model of diet-induced obesity (DIO) with subsequent weight reduction achieved via low-fat diet (LFD) feeding.ResultsEighteen C57BL/6NCrl male mice were divided into three subgroups: (1) control, mice were fed a standard AIN-76A (fat: 11.5 kcal %) diet for 12 weeks; (2) DIO, mice were fed a 58 kcal % high-fat diet (HFD) for 12 weeks; and (3) DIO + LFD, mice were fed a HFD for 8 weeks to induce obesity and then switched to a 10.5 kcal % LFD for 4 weeks. A switch to LFD feeding led to decreases in body weight, adiposity, and blood glucose levels in DIO mice. Microarray analysis of miRNA using The Mouse & Rat miRNA OneArray® v4 system revealed significant alterations in the expression of miRNAs in DIO and DIO + LFD mice. Notably, 23 circulating miRNAs (mmu-miR-16, mmu-let-7i, mmu-miR-26a, mmu-miR-17, mmu-miR-107, mmu-miR-195, mmu-miR-20a, mmu-miR-25, mmu-miR-15b, mmu-miR-15a, mmu-let-7b, mmu-let-7a, mmu-let-7c, mmu-miR-103, mmu-let-7f, mmu-miR-106a, mmu-miR-106b, mmu-miR-93, mmu-miR-23b, mmu-miR-21, mmu-miR-30b, mmu-miR-221, and mmu-miR-19b) were significantly downregulated in DIO mice but upregulated in DIO + LFD mice. Target prediction and function annotation of associated genes revealed that these genes were predominantly involved in metabolic, insulin signaling, and adipocytokine signaling pathways that directly link the pathophysiological changes associated with obesity and weight reduction.ConclusionsThese results imply that obesity-related reductions in the expression of circulating miRNAs could be reversed through changes in metabolism associated with weight reduction achieved through LFD feeding.

Far‐Infrared Radiation Promotes Angiogenesis in Human Microvascular Endothelial Cells via Extracellular Signal‐Regulated Kinase Activation

This study was designed to determine the in vitro angiogenic ability of far‐infrared (FIR) radiation in the skin‐derived cultured human microvascular endothelial cells and to elucidate the role of mitogen‐activated protein kinases (MAPKs) in this process. The results revealed that FIR radiation from a WSTM TY301 FIR emitter activated p38 and extracellular signal‐regulated kinase (ERK), but not Akt or c‐Jun N‐terminal protein kinases (JNK), and significantly promoted angiogenesis by increasing tube formation in Matrigel and the migration of cells across an eight micron polyester filter. The addition of 50 μm PD98059, a MEK inhibitor, significantly inhibited the activation of ERK and the enhanced angiogenesis; in contrast, the inhibition of p38 phosphorylation did not inhibit the enhanced angiogenesis. After FIR radiation, there was no increase in vascular endothelial growth factor (VEGF) isoforms (VEGF‐A, ‐B, ‐C and ‐D) mRNA and VEGF protein, no increase phosphorylation of endothelial nitric oxide synthase (eNOS) detected using Western blotting, and no increase in NO production detected using flow cytometry in cells pre‐incubated with the cell‐permeable NO‐binding dye diluted 4‐amino‐5‐methylamino‐2′, 7′‐difluorofluorescein diacetate (DAF‐FM DA). This study revealed that FIR radiation possesses in vitro angiogenic activity via the activation of the MEK/ERK but not the VEGF/Akt/eNOS‐dependent signaling pathways.