Non-invasive three-dimensional bone-vessel image fusion using black bone MRI based on FIESTA-C.

AIM: To evaluate the image quality of bone-vessel fused volume-rendering (VR) images reconstructed by three-dimensional "black bone" magnetic resonance imaging (MRI) based on the fast imaging employing steady-state acquisition cycled phases (FIESTA-C) sequence and time-of-flight magnetic resonance angiography (TOF-MRA). MATERIALS AND METHODS: Seventeen patients were analysed in this retrospective study. All patients underwent both MRI techniques including FIESTA-C and TOF-MRA and computed tomography angiography (CTA). MRI- and CT-based bone-vessel VR images were reconstructed. Visual depictions of frontal and parietal branches from the superficial temporal artery (STA) were independently scored by three experienced radiological technologists using a four-grade system. RESULTS: In the visual evaluation, the scores of the both right and left frontal branches in MRI-based VR image were significantly larger those at CT (p<0.01, respectively). The scores of both the right and left parietal branches tended to be larger in MRI-based than that in CT-based VR imaging, but were not significantly so (p=0.06, 0.13 respectively). In the interobserver agreement analysis, κ values were all good (range: 0.6-0.76) for STA branch evaluation in MRI-based VR images. CONCLUSION: MRI bone-vessel fused VR imaging can non-invasively depict STA frontal branches with better visibility compared to the CT-based VR imaging. This technique may be useful for the preoperative evaluation of donor branches for STA-middle cerebral artery bypass surgery.

Calcium influx through the TRPV1 channel of endothelial cells (ECs) correlates with a stronger adhesion between monocytes and ECs.

PURPOSE: Atherosclerosis is thought to be initiated by the transendothelial migration of monocytes. In the early stage of this process, the adhesion of monocytes to endothelial cells is supported by an increase in the intracellular concentration of calcium ion ([Ca(2+)]i) in endothelial cells. However, the main source of Ca(2+) has been unclear. In this study, the changes in ionic transmittance and [Ca(2+)]i due to the adhesion of monocytes were continuously measured by an electrophysiological technique and fluorescent imaging. Especially, we focused on transient receptor potential vanilloid channel 1 (TRPV1) as a Ca(2+) channel that could influence the adhesion of monocytes. MATERIAL AND METHODS: Whole-cell current was continuously recorded in human umbilical vein endothelial cells (HUVECs) by a patch electrode. RESULTS: The adhesion of monocytes (THP-1) induced a transient inward current in HUVECs, as well as an elevation of [Ca(2+)]i. This inward element was abolished by the application of 100 nM SB366,791, a selective antagonist of TRPV1 channel. Furthermore, SB366,791 significantly decreased the number of THP-1 cells that adhered to HUVECs (control: 231 ± 38, SB366,791: 96 ± 16 cells/mm2). CONCLUSION: These results suggest that an inward calcium current via the TRPV1 channels of endothelial cells correlates with a stronger adhesion between monocytes and endothelial cells.

Antihypertensive effect of a fixed-dose combination of losartan/hydrochlorothiazide in patients with uncontrolled hypertension: a multicenter study.

BACKGROUND: Achieving adequate blood pressure (BP) control often requires more than one antihypertensive agent. The purpose of this study was to determine whether a fixed-dose formulation of losartan (LOS) plus hydrochlorothiazide (HCTZ) (LOS/HCTZ) is effective in achieving a greater BP lowering in patients with uncontrolled hypertension. METHODS: The study was a prospective, multicenter, observational trial exploring the antihypertensive effect of a single tablet of LOS 50 mg/HCTZ 12.5 mg. A total of 228 patients whose BP had previously been treated with more than one antihypertensive agents without having achieved BP goal below 130/80 mmHg enrolled in the study. RESULTS: A significant decrease in systolic and diastolic BP was observed in both clinic and home measurement after switching from the previous treatment to LOS/HCTZ. There was a significant decrease in both B-type natriuretic peptide (BNP) and urinary albumin creatinine (Cr) excretion ratio (ACR), especially in patients with elevated values. In contrast, there was a significant increase in serum Cr concentration in conjunction with a decrease in estimated glomerular filtration rate (eGFR). Overall serum uric acid (UA) concentration increased, whereas in patients with hyperuricemia there was a significant reduction in this value. CONCLUSION: Switching to LOS/HCTZ provides a greater reduction in clinic and home BP in patients with uncontrolled hypertension. This combination therapy may lead to cardio-, reno protection and improve UA metabolism.

Action of neuwiedase, a metalloproteinase isolated from bothrops neuwiedi venom, on skeletal muscle: an ultrastructural and immunocytochemistry study

The damaging effects of neuwiedase, a non-hemorrhagic snake venom metalloproteinase from P-I class, on gastrocnemius muscle are studied herein. Following neuwiedase injection, ultrastructural alterations were detected early showing disarrangement of skeletal muscle fibers (characterized by discontinuity of Z lines), mitochondrial swelling, and disruption of plasma membrane and basal lamina. Degradation of skeletal muscle and the appearance of an amorphous substance, primarily composed of cellular debris, were noted after 24 hours. The presence of neuwiedase at the injection site (detected by immunocytochemistry) revealed highly specific labeling of myofibril components of damaged myocytes. In addition, proteolysis of muscle proteins assayed through myofibrils extracted from gastrocnemius muscle indicated that neuwiedase provoked degradation of myofibrils, especially myosin. These results suggest that skeletal muscle damage, induced by neuwiedase, is probably due to its proteolytic action on myofibrils, which are responsible for the maintenance of the cellular architecture.

Lipid microsphere formulation containing rifampicin targets alveolar macrophages.

The study demonstrated that lipid microspheres (LM) containing rifampicin (LM-RFP) could deliver the drug to alveolar macrophages in vitro and in vivo, and that intranasal administration to animals could achieve preferential accumulation in the lungs with less effect on the liver. The LM-RFP particles had a mean diameter of 247.2 +/- 75.7 nm, and their size remained stable when stored at 4 degrees C or 25 degrees C for at least 4 weeks. In vitro uptake of [(3)H]LM-RFP by alveolar macrophages was over 4 times higher than that of unencapsulated [(3)H]RFP, whereas the in vivo uptake was 30 times higher. Flow cytometric analysis and confocal laser scanning microscopy confirmed that LM could deliver the encapsulated drug effectively to alveolar macrophages in vitro and in vivo. Intranasal administration of [(3)H]LM-RFP to normal mice resulted in preferential pulmonary uptake of the drug and lower levels in the blood and liver compared with administration of unencapsulated [(3)H]RFP. In conclusion, LM-RFP could be a promising preparation for delivery via the respiratory tract to tuberculosis (TB) and TB/HIV patients.

Interaction of cancer cells with platelets mediated by Necl-5/poliovirus receptor enhances cancer cell metastasis to the lungs.

Necl-5 is an immunoglobulin (Ig)-like molecule that was originally identified as a poliovirus receptor and is often upregulated in cancer cells. We recently found that it colocalizes with integrin alpha(v)beta(3) at the leading edges of moving cells and enhances growth factor-induced cell movement and proliferation. Upon cell-cell contact, Necl-5 is removed from the cell surface by its trans-interaction with the cell adhesion molecule nectin-3, resulting in reduced cell movement and proliferation. Here, we investigated the role of Necl-5 in the interaction of cancer cells with platelets. Necl-5 was upregulated in CT26 cells, a colon adenocarcinoma cell line. When CT26 cells were injected into the tail vein of mice, they were arrested in the pulmonary vessels by adhering to platelets and subsequently metastasized to the lungs. Overexpression of Necl-5 in CT26 cells enhanced this metastasis, while inhibition of the trans-interaction of Necl-5 with CD226 by an anti-Necl-5 monoclonal antibody reduced the metastasis. Depletion of platelets by treatment with a rabbit anti-mouse platelet serum reduced the Necl-5-enhanced metastasis in mice. Thus, the trans-interaction of upregulated Necl-5 in cancer cells with its counter-receptor in platelets, probably CD226, is critical for efficient metastasis of cancer cells to the lungs.

Long-term effect of intravenous immunoglobulin on anti-MuSK antibody-positive myasthenia gravis.

Anti-muscle-specific receptor tyrosine kinase (MuSK) antibody-positive myasthenia gravis (MG) patients show various responses to conventional immunosuppressive treatment and some patients are resistant to these therapies. We report a 50-year-old Japanese man with anti-MuSK antibody-positive MG, who showed no or poor response to various therapies, including plasmapheresis, corticosteroid, and tacrolimus. The patient was then treated with intravenous immunoglobulin (IVIG), and showed a good response that persisted over 20 months. The outcome of this case suggests that IVIG treatment may be an effective therapeutic option for anti-MuSK antibody-positive MG, with a potentially long-term effect.

Purification and functional characterization of two fibrinogenolytic enzymes from Bothrops alternatus venom

Two fibrinogenolytic enzymes, Bothrops alternatus metalloprotease isoform (BaltMP)-I and II, were purified from Bothrops alternatus venom using Diethylaminoethyl (DEAE) Sephacel, Sephadex G-75 and Heparin-Agarose column chromatography. Purified BaltMP-I and II ran as single protein bands on analytical polyacrylamide gel electrophoresis and showed molecular weights of 29000 and 36000, respectively, under reducing conditions in sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). BaltMP-II, but not BaltMP-I, displayed blood-clotting activity in bovine plasma, which was about 10-fold higher than that of the crude venom. Both enzymes were proteolytically active against bovine fibrinogen as substrate. When fibrinogen and each enzyme were incubated at 37°C, at a ratio of 1:100 (w/w), BaltMP-II cleaved preferentially the Aalpha -chain and more slowly the Bbeta -chain. The action of BaltMP-I was similar, but lower. None of the proteases degraded the gamma-chain of fibrinogen. The fibrinogenolytic activity of the enzymes was inhibited by 1,10-phenanthroline, suggesting they are metalloproteases. Since both enzymes were found to cause defibrinogenation when intraperitoneally (i.p.) administered to mice, they can be of medical interest as a therapeutic agent in the treatment and prevention of arterial thrombosis.(AU)

Neutralization of some hematological and hemostatic alterations induced by neuwiedase, a metalloproteinase isolated from Bothrops neuwiedi pauloensis snake venom, by the aqueous extract from Casearia mariquitensis (Flacourtiaceae).

The aqueous extract from the leaves of Casearia mariquitensis (C. m.), a plant found in Brazilian open pastures, was assayed for its ability to inhibit some hematological and hemostatic effects induced by neuwiedase, a 22 kDa class P-I metalloproteinase from the venom of the South American pit viper Bothrops neuwiedi pauloensis. The aqueous extract from C. m. was able to neutralize the hematological alterations induced by the crude venom (C.V.) upon erythrocytes when the venom was incubated at a ratio of 1:10 (w/w, venom/extract), but it did not neutralize the platelet decreasing ability of C.V. The plasma fibrinogen concentration decreased approximately 36% and 83% when 0.6 LD(50) of the C.V. or neuwiedase, respectively, were injected by i.p. route in mice, and the aqueous extract from C. m. was able to inhibit this effect. The Bbeta fibrinogen chain was protected against degradation caused by crude venom and neuwiedase when the venom or toxin were incubated with C. m. extract. We also observed that this extract exerted a very slight effect on the clotting time, prolonging it only to a little extent. The pulmonary hemorrhage induced by neuwiedase when injected intravenously with 0.6 LD(50) was completely inhibited when this toxin was incubated with the extract at a ratio of 1:10 (w/w, toxin/extract). It is concluded that C. m. displays components able to inhibit some hematological and systemic alterations induced by C.V.

[A study on the mechanism of enhanced diuresis following direct hemoperfusion with polymyxin B-immobilized fiber].

In the present study, we applied direct hemoperfusion with polymyxin B-immobilized fiber(PMX-DHP) to patients who developed endotoxin shock after laparotomy, and examined the influence of PMX-DHP on the kidney function. Seven patients were enrolled in this study, whose conditions were matched to the following criteria: 1) endotoxin shock was highly suspected, 2) blood pressure became stable before PMX-DHP was indicated, 3) renal function(demonstrated with creatinine clearance(CCr) and fractional excretion of sodium (FENa)) was proven before the surgery. All patients underwent emergency surgery in Fuji City General Hospital because of perforative peritonitis. A 2-hour session of PMX-DHP was performed on the day of the laparotomy and the second 2-hour treatment was performed the following day. Urine was collected at 2 hours before starting PMX, during the treatment, and 2 hours after PMX-DHP, and urine volume(U-Vol), sodium and creatinine levels of urine were monitored. Sodium and creatinine levels in the serum were measured at the start and end of the PMX-DHP session. Average atrial natriuretic polypeptide (ANP) was obtained using a total of 8 samples from the 14 treatment sessions. Parameters of hemodynamics such as pulmonary capillary wedge pressure(PCWP) were monitored at the start and end of PMX-DHP session. Urine volume increased significantly during and after PMX-DHP. The change in urine volume correlated significantly with the change in CCr during PMX-DHP, and with the change in FENa after PMX-DHP. The change in FENa was significantly correlated with the changes in hemodynamic factors such as PCWP and with the change in serum ANP, but no significant correlation was observed between the change of CCr and the other parameters. In conclusion, the early increase in urine volume with PMX-DHP treatment might be attributable to the increase in glomerular filtration independently of systemic hemodynamic factors.

[Clinical evaluation of cases with massive ascites accumulation after discontinuation of CAPD: an attempt to establish the concept of the pre-sclerosing encapsulating peritonitis(pre-SEP) state].

Sclerosing encapsulating peritonitis(SEP) is a most serious complication of continuous ambulatory peritoneal dialysis(CAPD). Although the criteria of diagnosis and guidelines for therapy of SEP have been proposed by the Japanese SEP Study Group already, SEP is refractory to treatment when the disease process is complete. It is important to detect the latent phase of SEP(pre-SEP state) in order to treat patients at an early stage. We evaluated the characteristics of ascites in four patients with massive ascites accumulation after discontinuation of CAPD. Age and the duration of CAPD of the subjects were 53.3 +/- 9.7 years and 126.5 +/- 6.8 months, respectively. However, the patients were withdrawn from CAPD because of peritonitis or ultrafiltration failure. We also followed cytokines and parameters of collagen metabolism of ascites in two patients during adrenocorticosteroid therapy and conducted a histopathological evaluation of the peritoneum of an autopsy case who had died of pneumonia. Ascites seems to be exudative because of the high concentration of protein, cytokines and parameters of collagen metabolism such as interleukin-1 beta, interleukin-6, transforming growth factor-beta 1, procollagen 3 peptide, and type IV collagen 7S, the levels of which were 21.3 +/- 9.3 pg/ml, 8,153 +/- 7,327 pg/ml, 6.7 +/- 3.6 ng/ml, 89.3 +/- 67.8 U/ml, and 59.0 +/- 36.2 ng/ml, respectively. The histopathological findings of the peritoneum from the autopsy case showed dense fibrous tissue permeated with inflammatory infiltration and widespread infiltration of fibrin. These findings suggested that the peritoneum was inflamed when massive ascites accumulated. The amount of ascites and concentration of cytokines and parameters of collagen metabolism of ascites diminished during adrenocorticosteroid therapy. We concluded that massive and refractory accumulation of ascites appearing after the discontinuation of CAPD should be regarded as a sign of the pre-SEP state, and prophylactic treatment should be started at this stage of disease.

Effects of aqueous extract of Casearia sylvestris (Flacourtiaceae) on actions of snake and bee venoms and on activity of phospholipases A2.

The crude aqueous extract from the leaves of Casearia sylvestris, a plant found in Brazilian open pastures, was assayed for its ability to inhibit phospholipase A2 (PLA2) activity and some biological activities of bee and several snake venoms, and of a number of isolated PLA2s. The extract induced partial inhibition of the PLA2 activity of venoms containing class I, II and III PLA2s. When tested against the purified toxins, it showed the highest efficacy against class II PLA2s from viperid venoms, being relatively ineffective against the class I PLA2 pseudexin. In addition, C. sylvestris extract significantly inhibited the myotoxic activity of four Bothrops crude venoms and nine purified myotoxic PLA2s, including Lys-49 and Asp-49 variants. The extract was able to inhibit the anticoagulant activity of several isolated PLA2s, with the exception of pseudexin. Moreover, it partially reduced the edema-inducing activity of B. moojeni and B. jararacussu venoms, as well as of myotoxins MjTX-II and BthTX-I. The extract also prolonged the survival time of mice injected with lethal doses of several snake venoms and neutralized the lethal effect induced by several purified PLA2 myotoxins. It is concluded that C. sylvestris constitutes a rich source of PLA2 inhibitors.

[Factors influencing residual renal function of CAPD patients].

It is generally accepted that residual renal function has significant meaning in dialysis adequacy of CAPD patients. However, the factors influencing the residual renal function have not been investigated yet. We evaluated the consequences of following factors on residual urine volume in 50 CAPD patients: deterioration rate of renal function (slope of I/serum creatinine) before dialysis, renal creatinine clearance at the initial point of dialysis and the episodes of rapid reduction of residual urine volume. There was no correlation between the deterioration rate of renal function before dialysis and the residual urine volume. On the contrary, there was significant correlation between renal creatinine clearance at the initial point of dialysis and the residual urine volume in the first 4 years on CAPD. We recognized 67 episodes of rapid reduction of residual urine volume after initiation of CAPD. Inappropriate management or complications attributed to the rapid urine volume reduction in 50 episodes. The residual urine volume did not recover to the previous level in 40 episodes. We concluded that early initiation of CAPD and evasion of decreasing residual urine volume caused by inappropriate management or complications have important meaning and will preserve residual renal function of CAPD patients.

Orthotopic ileal neobladder in male patients: functional outcomes of 66 cases.

BACKGROUND: Orthotopic urinary diversion has become the preferred form of bladder reconstruction after cystectomy. We report on our experience with 66 male patients undergoing this procedure from November 1990 to February 1998. METHODS: A neobladder was constructed using an ileal segment with a Hautmann type bladder. Complications were assessed and subdivided into early and late types. Voiding function was evaluated in terms of voiding pattern and continence. Median follow up was 19.5 (range 3.5-87.7) months. RESULTS: There was one (1.5%) perioperative death. The most frequent pouch-related and unrelated early complications were persistent urine leak (7.6%) and prolonged ileus (16.7%), respectively, the majority of cases of which were managed conservatively. Analysis of late complications revealed 6.2% ureteroileal stenosis and 1.5% urethrointestinal stenosis rates, but no case of bladder stone formation. Of the 61 patients in whom voiding function was evaluable, 95.1% achieved excellent daytime continence, while only 67.2% had night-time continence. With regard to posture at voiding, 23 (37.7%) voided in a sitting position. Three of the patients (4.9%) were unable to void and required regular intermittent catheterization. CONCLUSIONS: An orthotopic neobladder can be constructed with acceptable morbidity and excellent functional results. We believe that orthotopic urinary diversion offers an attractive alternative to a bladder substitute when cystectomy is required.

Purification and partial characterization of a new proteolytic enzyme from the venom of Bothrops moojeni (CAISSACA).

A basic serine protease which is active on casein and fibrinogen was purified from Bothrops moojeni venom using a single step chromatography on a CM-Sepharose fast flow column. The enzyme, MOO3, was not hemorrhagic and presented only a trace of blood-clotting activity. Synthetic chromogenic substrates (azoacasein and azoalbumin) where not hydrolyzed by MOO3. Using polyacrylamide gel electrophoresis at pH 4.3, MOO3 showed as a single protein band. Using sodium dodecyl sulfate-polyacrylamide electrophoresis, MOO3 behaved as a single-chain protein with an approximate mol. weight of 27,000, both in the presence and absence of beta-mercaptoethanol. Its pI was 7.8 by electrofocusing. The enzyme did not contain neutral carbohydrates and its N-terminal amino acid was alanine. The amino acid composition showed 249 residues/mole, a high content of hydrophilic amino acids and 14 half-cystine residues, which should account for 7 disulfide bonds. The protease cleaved the A-alpha chain faster than the B-beta of bovine fibrinogen and showed no effect on the delta-chain. Specific esterolytic activity of MOO3 on alpha-N-tosyl-l-arginine methyl ester was 29.64 mumol min-1 x mg-1. MOO3 represented 1.42% (w/w) of the initial desiccated venom. Its proteolytic activity was inhibited by beta-mercaptoethanol, leupeptin, phenylmethylsulphonyl fluoride and ethylenediamine tetraacetate.

Voiding function of orthotopic ileal neobladder in women.

OBJECTIVES: Although the advent of orthotopic lower urinary reconstruction in women is a major achievement in the evolution of urinary diversion, the mechanism of voiding dysfunction remains incompletely understood. We report on the voiding function of ileal neobladder in 12 female patients. METHODS: A neobladder was constructed using an ileal segment. Voiding function was evaluated in terms of voiding pattern and continence. Chain cystography was performed postoperatively to detect the existence of urethral angulation. Median follow-up was 33.2 months (range 8.4 to 77.4). RESULTS: Of the 1 2 patients, 10 (83.3%) achieved excellent daytime continence, and 6 (50%) had nighttime incontinence despite regular voiding during the night. Three patients (25%) sometimes or often performed self-catheterization because of difficulty in urinating. One patient (8.3%) was unable to void and required regular intermittent catheterization. Chain cystogram revealed urethral angulation in the 3 patients with difficulty in urinating or hypercontinence. CONCLUSIONS: An orthotopic neobladder can be constructed in women with excellent functional results. Urethral angulation appears to be one of the main mechanisms for voiding dysfunction, and further studies on the functional anatomy of the female urethra are needed to improve the voiding function of the orthotopic neobladder in women.