Biomedical subjects

Jian Tu

Publications and source records attributed to Jian Tu.

12 recordsLinked to original sources

PubMed2006

Responses of arteriovenous malformations to radiosurgery: ultrastructural changes.

OBJECTIVE: To examine the ultrastructural changes in arteriovenous malformations (AVMs) after radiosurgery and to explore the possible mechanisms of posttreatment obliteration and hemorrhage. METHODS: Twenty-two specimens, among them three irradiated AVMs (size, 3-6 cm), 15 nonirradiated AVMs, and four normal controls were processed for ultrastructural study immediately after removal. Transmission electron microscopy was used to compare the vasculature of irradiated AVMs with nonirradiated AVMs and normal controls. RESULTS: Thirty-three months postradiosurgery, partial vaso-occlusion (36-74% lumen) occurred by coagulation of cytoplasmic debris and proteinaceous material leaking from the endothelium. Forty-eight months postradiosurgery, heterogeneous thrombus formation (86-96% lumen) with fibrinoid and proteinaceous materials was observed. Sixty-four months postradiosurgery, complete luminal closure (90-100% lumen) by a fibrin thrombus was seen in vessels with diameters up to 5.5 mm including feeding arteries and draining veins. In occluded vessels, there was extensive degeneration of endothelial cells, subendothelial fibroblasts, and myofibroblasts. Neoproliferation and endothelialization of smooth muscle cells with Weibel-Palade bodies was observed in arteries. CONCLUSION: Radiosurgery causes irreversible cellular damage of the vascular wall. Partial vaso-occlusion that increases blood flow in remaining vessels and degenerative changes on the blood-brain barrier may contribute to hemorrhage at early stage postradiosurgery. Radiosurgery stimulates neoproliferating and endothelializing smooth muscle cells in vessel walls, which might lead to narrowing of the vessel lumina. Complete vaso-occlusion achieved 64 months postradiosurgery suggested a minimum follow-up duration of 5 years to determine final outcome of radiosurgery. Histological end point of vaso-occlusion of AVMs takes longer time than neuroimaging endpoint of complete obliteration.

Adolescent↗

PubMed2006

Ultrastructure of perinidal capillaries in cerebral arteriovenous malformations.

OBJECTIVE: The ultrastructure of perinidal capillaries in cerebral arteriovenous malformations (AVMs) was examined to clarify their pathomorphological features. METHODS: Fifteen AVM specimens were dissected and divided into perinidal and intranidal groups and processed for ultrastructural study immediately after surgical removal. Eleven of the patients had presented with hemorrhage. Tissue from four normal controls was also studied. Electron microscopy was used to compare features of the blood-brain barrier and endothelial cells (ECs) of capillaries in perinidal, intranidal, and controls. RESULTS: Perinidal capillaries demonstrated abnormal ultrastructure of the blood-brain barrier with no basement membranes and astrocytic foot processes. ECs had fenestrated luminal surfaces. Large gaps were observed at endothelial intercellular junctions. ECs contained numerous filopodia, large numbers of cytoplasmic processes, numerous micropinocytotic vesicles, and the cytoplasm contained more filaments than those observed in controls. Pericytes were rich in pinocytotic vesicles, vacuoles, and filaments. Their processes were in close contact with ECs. Weibel-Palade bodies were present in perinidal ECs. CONCLUSION: The absence of blood-brain barrier components in perinidal capillaries may contribute to extravasation of red blood cells into the surrounding brain in the absence of major hemorrhage and explain the gliosis and hemosiderin occasionally seen around AVMs. Cellular differentiation and proliferation in perinidal capillaries should be included in a systematic study aimed at a better understanding of the mechanisms underlying the recurrence of surgically removed AVMs.

Adolescent↗

PubMed2006

[A reverse transcriptase-polymerase chain reaction method improving the specific amplification of caveolin-1 gene sequences].

OBJECTIVE: To construct a reverse transcriptase-polymerase chain reaction (RT-PCR)approach that can improve the specificity of primers while dropping down the nonspecific amplification. METHODS: In the recent study we reported a new RT-PCR assay which improved markedly the specificity. However its efficiency of regressing nonspecific amplification remains to be accurately checked and further documented. In primer design, we looked over again some sequences that showed differences at 5' or 3' ends between human CAV1 and mouse Cav1 genes. cDNAs and the diluted plasmids which harbored the sequence of human CAV1 or mouse Cav1 gene were chosen as the templates. The ordinary PCR compared with one, of which primers modified by phosphorothioate and combined with proofreading polymerase, for their efficiencies of nonspecific amplification inhibited. RESULTS: Taq DNA polymerase without proofreading activity could efficiently catalyze the extension of primers with a single or multiple mismatched base pairs at the 3' terminus, but the kind of primer extension can be effectively blocked by phosphorothioate modified primers combined with proofreading polymerase. Compared with ordinary PCR reaction, this new PCR method can effectively regress the primer mismatched amplification of 50 ng DNA almost equaling to 2 x 10(4) unmatched template copies in a final volume of 50 microL. CONCLUSION: Compared with the first generation of polymerases with or without proofreading activities mediating RT-PCR reaction, the introduction of nuclease-resistant 3' modified primers (3' phosphorothioate primer extension) can offer more simplicity, accuracy, and also decrease cost.

Animals↗

PubMed2005

Estimation of logP(ow) values for neutral and basic compounds by microchip microemulsion electrokinetic chromatography with indirect fluorimetric detection (muMEEKC-IFD).

Microchip microemulsion electrokinetic chromatography with indirect fluorimetric detection (muMEEKC-IFD) was used to obtain logP octanol/water (logP(ow)) values for neutral and basic compounds. Six compounds, with logP(ow) values between 0.38 and 5.03, were used to create a calibration curve relating the log of retention factors (logk) obtained from muMEEKC-IFD with the known logP(ow) values. The logP(ow) values for six additional compounds were determined using the logk values obtained by muMEEKC-IFD and the linear relationship between logP(ow) and logk established for the standard compounds. The muMEEKC-IFD buffer was composed of 50 mM 3-[cyclohexylamino]-1-propane-sulfonic acid (CAPS) buffer (pH 10.4) containing 1.2% n-heptane (v/v), 2% sodium dodecylsulfate (w/v), 8% 1-butanol (v/v) and 4 microM 5-carboxytetramethyl-rhodamine (TAMRA) as the fluorophore probe for indirect detection. The muMEEKC-IFD provided an accurate method for estimating logP(ow) values and also a means for analyzing compounds that are non-fluorescent.

Calibration↗

PubMed2005

Ultrastructural characteristics of hemorrhagic, nonhemorrhagic, and recurrent cavernous malformations.

OBJECT: Ultrastructural characteristics of hemorrhagic, nonhemorrhagic, primary, and recurrent central nervous system cavernous malformations (CMs) were examined in an attempt to clarify their pathological mechanisms. METHODS: Thirteen specimens (nine from samples of CMs and four from healthy control tissue) were processed for ultrastructural study immediately after surgical or postmortem removal, by fixation in glutaraldehyde/formalin and postfixation in OsO4. Transmission electron microscopy was used to examine the vascular walls, endothelium, subendothelium, and cytoplasmic organelles. The vascular walls in CMs demonstrated abnormal ultrastructure with no basement membranes and astrocytic foot processes. Pericytes were rarely seen. Single-layer lining endothelial cells showed fenestrated luminal surfaces. Large gaps were observed at intercellular junctions between endothelial cells, and large vesicles with extremely thin plasma membranes bordering the lumen were common in the lesions that had previously hemorrhaged. Endothelial cells of recurrent CMs had more Weibel-Palade bodies, filopodia, cytoplasmic processes, micropinocytotic vesicles, and filaments than those in primary lesions and normal control tissues. CONCLUSIONS: The absence of the blood-brain barrier, normal supporting wall structure, and large vesicles bordering the lumen of CM vessels may explain leakage of red blood cells into surrounding brain in the absence of major hemorrhage. Proliferation of residual abnormal endothelial cells may contribute to the recurrence of surgically removed CMs.

Adolescent↗

PubMed2004

Comparison of fetal porcine aggregates of purified beta-cells versus islet-like cell clusters as a treatment of diabetes.

Fetal pig islet-like cell clusters (ICCs) have the potential to reverse diabetes 1-5 months after transplantation. In a fetal ICC, however, beta-cells constitute only 6-8% of the cells, in contrast to 65% in an adult pig islet. Attempts to purify fetal beta-cells from cell clusters and compare their function to that of ICCs have not been shown previously. Beta-cells were purified from ICCs isolated from the fetal pig pancreas. These were then aggregated and maintained in culture for 3 days. ICCs were isolated from fetal pig pancreas and allowed to round up in culture for 3 days. Transplantation of aggregates and ICCs (10,000 and 12,600, respectively) into diabetic immunoincompetent mice resulted in normoglycemia at 18 +/- 2 and 8 +/- 1 weeks, respectively (p = 0.0006). Removal of grafts after normalization of blood glucose levels resulted in rapid return of hyperglycemia in both groups. In conclusion, a purified population of immature beta-cells can be produced from the fetal pig pancreas. The reason these cells take longer than ICCs to reverse diabetes when transplanted is postulated to be because of the relative lack of precursor cells from which beta-cells differentiate. This finding may have implications for stem cell therapy, as other cell types, other than purified beta-cells, may be necessary for appropriate function in vivo.

Animals↗

PubMed2003

Altered subarachnoid space compliance and fluid flow in an animal model of posttraumatic syringomyelia.

STUDY DESIGN: A histologic study of cerebrospinal fluid tracers in Sprague-Dawley rats undergoing lumboperitoneal shunt insertion in the excitotoxic animal model of posttraumatic syringomyelia (PTS). OBJECTIVES: To determine the effects of cerebrospinal fluid (CSF) diversion from the subarachnoid space on perivascular flow (PVS) and syrinx formation in posttraumatic syringomyelia. SUMMARY OF BACKGROUND DATA: In an animal model of PTS, fluid enters syringes from the subarachnoid space via perivascular spaces. Preferential PVS flow occurs at the level of the syrinx. It has been suggested that arachnoiditis predisposes to posttraumatic syringomyelia formation by obstructing subarachnoid cerebrospinal fluid flow and enhancing perivascular flow. MATERIALS AND METHODS: Thirty-two male Sprague-Dawley rats were investigated using the CSF tracer horseradish peroxidase (HRP), the excitotoxic model of PTS, and lumboperitoneal shunt insertion. Five experimental groups consisted of normal controls, syrinx only and shunt only controls, and shunt insertion before or after syrinx formation. In all groups except normal controls, CSF flow studies were performed 6 weeks after the final intervention. Grading scales were used to quantify HRP staining. RESULTS: All excitotoxic model animals formed syringes. Perivascular flow was greatest at the level of the syrinx. Cerebral cortex perivascular flow was significantly reduced after shunt insertion in animals with a syrinx (P < 0.05). Shunt insertion did not alter syrinx length or size. There were no significant differences between shunt and syrinx first groups. CONCLUSIONS: Increasing caudal subarachnoid space compliance with a shunt does not affect local CSF flow into the spinal cord and syrinx. These results suggest that localized alterations in compliance, as opposed to obstruction from traumatic arachnoiditis, may act as an important factor in syrinx pathogenesis.

Animals↗

PubMed2003

Application of multiplexed capillary electrophoresis with laser-induced fluorescence (MCE-LIF) detection for the rapid measurement of endogenous extracellular signal-regulated protein kinase (ERK) levels in cell extracts.

Multiplexed (96-lane) capillary electrophoresis with laser-induced fluorescence (MCE-LIF) detection was used for the rapid analysis of extracellular signal-regulated protein kinase (ERK) levels from in vitro cell extracts. The levels of ERK enzyme in cell extracts were determined by monitoring the conversion of a fluorescent-labeled peptide substrate to a phosphorylated fluorescent-labeled peptide product using MCE-LIF. The incorporation of a fluorescent internal standard was found to improve the precision of the analysis. The enzyme assay conditions including substrate concentration, reaction time and enzyme linear range were rapidly optimized using the MCE-LIF approach for both direct and immunoprecipitation-based ERK assays. The levels of ERK from in vitro cell extracts stimulated with angiopoietin 1 (Ang1*) were determined using the MCE-LIF approach. The advantages of MCE-LIF for developing and applying enzyme assays, as well as the figures of merit for the direct and immunoprecipitation ERK assays, are discussed.

Cell Extracts↗

PubMed2003

Determination of endogenous extracellular signal-regulated protein kinase by microchip capillary electrophoresis.

The application of microchip capillary electrophoresis (CE) to the assay of extracellular signal-regulated protein kinase (ERK) is presented. In this assay, ERK catalyzes the transfer of gamma-phosphate from adenosine 5(')-triphosphate to the threonine residue of a fluorescently labeled nonapeptide (APRTPGGRR), and the phosphorylated and nonphosphorylated peptides were detected by fluorescence. The phosphorylated and nonphosphorylated peptides and the internal standard were separated within 20s, and the increase in magnitude of the phosphorylated peptide peak was monitored to assess ERK activity. ERK reactions were prepared off-chip and analyzed on a single-lane glass microchip fabricated by standard methods. It was demonstrated that microchip CE could be used to measure endogenous amounts of ERK by spiking known concentrations of recombinant ERK2 into the lysates of serum-starved human umbilical vein endothelial cells (HUVEC) and recovering between 90 and 100% for all samples. Endogenous ERK activity was determined by microchip where HUVEC were stimulated with 500pM vascular endothelial growth factor (VEGF) at different times before cell lysis. The results showed a transient VEGF-mediated ERK activation that peaked at 10min, which was consistent with previous reports using conventional techniques. The microchip assay provided a rapid, accurate, and precise alternative to conventional methods of determining endogenous ERK activity.

Cells, Cultured↗

PubMed2003

The role of excitotoxic injury in post-traumatic syringomyelia.

Fifty percent of patients with neurological deterioration from post-traumatic syringomyelia do not respond to treatment. Treatment failure is due in part to an incomplete understanding of the underlying aetiology. An animal model that mimics the human disease is required to investigate underlying pathophysiology and treatment options. A previous study was designed to mimic trauma-induced effects on the spinal cord that result in syringomyelia, combining an excitotoxic insult with kaolin-induced arachnoiditis. In this excitotoxic model, syringes were produced in 82% of animals. The aims of the current study were to improve the model to produce syringes in all animals treated, to examine the relative influences of excitotoxic injury and neuronal loss on syrinx formation, and to use magnetic resonance imaging (MRI) to examine syringes non-invasively. A temporal and dose profile of intraparenchymal quisqualic acid (QA) and subarachnoid kaolin was performed in Sprague Dawley rats. MRI was used to study four syrinx and six control animals. In one subgroup of animals surviving for 6 weeks, 100% (eight of eight) developed syringes. Syrinx formation and enlargement occurred in a dose and time dependent manner, whilst significant neuronal loss was only dose dependent. Animal syrinx histology closely resembled human post-traumatic syringomyelia. Axial T2-weighted MR images demonstrated syrinx presence. The results suggest that the formation of an initial cyst predisposes to syrinx formation in the presence of subarachnoid adhesions.

Animals↗

PubMed2002

Fluid flow in an animal model of post-traumatic syringomyelia.

More than a quarter of patients with spinal cord injury develop syringomyelia, often with progressive neurological deficit. Treatment options remain limited and long-term failure rates are high. The current poor understanding is impeding development of improved therapies. The source and route of fluid flow into syringes has been investigated using cerebrospinal fluid (CSF) tracers. Previous work using a model of canalicular syringomyelia has shown that fluid enters the dilated central canal from perivascular spaces. The aim of this study was to determine the source and route of fluid flow in an animal model of extracanalicular (post-traumatic) syringomyelia. A model of post-traumatic syringomyelia was established in 25 Sprague-Dawley rats with intraparenchymal injections of quisqualic acid and kaolin-induced arachnoiditis. Rats survived for 6 weeks before injection of the CSF tracer horseradish peroxidase into the cisterna magna. Examination of the spatial distribution of horseradish peroxidase at 0, 3, 5, 10, or 20 min after injection was used to determine the route of fluid flow. Horseradish peroxidase rapidly spread to the ventromedian fissure, perivascular spaces, central canal, and extracanalicular syrinx. Flow occurred into the syrinx prior to significant perivascular flow in the rostral spinal cord. Preferential flow into the syrinx occurred from the perivascular spaces of the central penetrating branches of the anterior spinal artery in the grey matter. Transparenchymal flow into the syrinx was less prominent than perivascular flow. This is the first report of fluid flow within the spinal cord in a model of post-traumatic syringomyelia. Fluid from perivascular spaces moves preferentially into extracanalicular syringes and the surrounding parenchyma. Obstruction to CSF flow and loss of compliance from traumatic arachnoiditis might potentiate fluid flow in the perivascular space.

Animals↗

PubMed2002

Role of NADH shuttles in glucose-induced insulin secretion from fetal beta-cells.

The NADH shuttle system, which transports the substrate for oxidative metabolism directly from the cytosol to the mitochondrial electron transport chain, has been shown to be essential for glucose-induced activation of mitochondrial metabolism and insulin secretion in adult beta-cells. We examined the role of these shuttles in the fetal beta-cell, which is immature in being unable to secrete insulin in response to glucose. The activity and concentration of the two key enzymes of the NADH shuttles, mitochondrial glycerol phosphate dehydrogenase (mGPDH) and mitochondrial malate dehydrogenase (mMDH), were eight- and threefold lower, respectively, in fetal compared with adult rat islets. Likewise, mGPDH and mMDH activity was fivefold lower in islet-like cell clusters (ICCs) and sevenfold lower in purified beta-cells compared with adult islets in the pig. The low level of enzyme activity was a result of low gene expression of the mitochondrial enzymes in the fetal beta-cells. Increasing NADH shuttle activity by transduction of fetal rat islets with mGPDH cDNA enabled the fetal islets to secrete insulin when stimulated with glucose. We concluded that the immaturity of the NADH shuttles contributes to the inability of fetal beta-cells to secrete insulin in response to glucose.

Adenoviridae↗