Selective depletion of major and minor histocompatibility antigen reactive T cells: towards prevention of acute graft-versus-host disease

Development of acute graft-versus-host disease (aGVHD) following HLA-identical sibling bone marrow transplantation (BMT) remains a serious complication. A selective depletion of T cells has proved to be effective in preventing aGVHD but is associated with relapse and increased incidence of infection. As aGVHD is directed mainly against epithelial tissues we examined whether it would be feasible to selectively deplete T cells reactive with epithelial cells whilst preserving other specificities. Donor T cells which express HLA-DR, CD25, CD69 and CD71 activation markers after cocultivation with patient keratinocytes were depleted using magnetic cell separation techniques. Depletion of major as well as minor histocompatibility antigen activated T cells revealed a significant (P = 0.004 and P = 0.031, respectively) 10-fold decrease in the frequency of donor T lymphocyte precursors reactive with patient keratinocytes. The frequency reactive with third-party and patient peripheral blood mononuclear cells, including leukaemia cells, remained unchanged, supporting the notion that aGVHD and graft-versus-leukaemia (GVL) may be separable. This alloantigen-specific depletion may be used in matched unrelated as well as HLA-identical sibling BMT for reducing aGVHD whilst conserving GVL.     

A risk-based approach to establish stability testing conditions for tropical countries

The external stability risk factors heat and moisture are evaluated with respect to the development of pharmaceutical products intended to be marketed in tropical and subtropical countries. The mean daily temperatures and dewpoints measured four times per day at selected places in Southeast Asia, South America, China, Southern Africa and the Caribbean are used to calculate the daily and monthly fluctuations of temperature and partial water vapour pressure, the mean kinetic temperature and the relative humidity. Based on these data, the hottest and the most humid place in each country or region are identified to reflect the worst case for the specific region. A formula to calculate safety margins for temperature and partial vapour pressure is introduced taking into consideration the difference between measured meteorological parameters and the stability testing conditions. An appropriate long-term stability testing condition is proposed for each selected country, related to the worst case for each specific region and the safety margins, as well as its classification in either Climatic Zone IVA or IVB.     

Bismuth biokinetics and kidney histopathology after bismuth overdose in rats

Bismuth induced nephrotoxicity has been reported to occur after acute overdoses of Bi-containing therapeutic drugs. We studied the development of bismuth induced nephropathy and bismuth biokinetics in rats. Bismuth nephropathy was induced in 33 young adult female Wistar rats weighing ca. 175 g by feeding them a single overdose of colloidal bismuth subcitrate containing 3.0 mmol Bi/kg at (t = 0). Control animals (n = 7) were fed the vehicle only. The animals were sacrificed after 1-48 h. Plasma creatinine increased from 51 +/- 6 micromol/l at t = 0 to 550 +/- 250 micromol/l after 48 h in the experimental group. The S3 segment of the proximal tubule showed epithelial cell vacuolation after 1 h and necrosis after 3 h. Cells of the S1/S2 segment demonstrated vacuolation after 6 h and necrosis after 12 h. Biokinetics of bismuth in blood could best be described with a one-compartment model characterized by an absorption half-life of 0.32 h and an elimination halflife of 16 h. The peak concentration of about 7.0 mg Bi/l was reached after 2 h. In conclusion, cells of the S3 segment of the proximal tubule necrotized first after an oral colloidal bismuth subcitrate overdose and biokinetics of Bi in blood was best described by a one-compartment model.     

Accumulation of Aluminium in Rat Liver: Association with Constituents of the Cytosol

Abstract: Aluminium (A1) accumulation occurs in the liver of renal patients and in patients on parenteral nutrition. Human hepatotoxicity is not proven. The role of the liver in storage and biotransformation of A1 and in development of osteo- and neurotoxicity is not clarified as yet. The aim of the present investigation was to study the storage of A1 in total liver and in subcellular liver fractions, and its association with soluble cytosolic molecular species. Therefore, rats were loaded with A1 prior to liver fractionation by ultracentrifugation, and equilibrium gel filtration chromatography of the cytosol, using a previously described method for A1 speciation in serum. A1 accumulated dose-dependently in liver and subcellular liver fractions, the lowest levels occurring in the cytosol. A dose-dependent elevation of A1 in the blood was also observed. Gelfiltration of the cytosol indicated that A1 was associated with a low molecular weight form which was not a citrate complex, and a high molecular weight form, which was larger than transferrin. No induction of and association with metallothionein occurred.     

Temozolomide in advanced malignant melanoma with small brain metastases

BACKGROUND: The efficacy of radiotherapy (RT) in patients who have brain metastases from melanoma is limited. In this study, the authors evaluated the efficacy of treatment with temozolomide in patients with metastatic melanoma, including small brain metastases, who did not require immediate RT and investigated the feasibility of deferring RT. METHODS: Patients with brain metastasis were identified from 3 prospective studies of temozolomide (with or without immunotherapy) for metastatic melanoma. Patients with brain metastasis that measured >2 cm, extensive edema, and localization in the brain stem were excluded from the study. For the current analysis, patients with leptomeningeal metastasis and patients who received previous stereotactic RT were excluded. In patients who achieved a systemic response or stabilization to temozolomide, the response of brain metastasis and the necessity for palliative cranial RT were evaluated. RESULTS: Among 179 patients who received temozolomide for advanced melanoma, 52 patients with brain metastasis were evaluable. Stabilization of systemic metastasis was noted in 7 of 52 patients (13%), and there were 6 responses (5 partial responses and 1 complete response; 11%); thus, in those 13 patients, 6 had stabilization of brain metastasis (11%) and 5 had a response (2 partial responses and 3 complete responses; 9%). Immunotherapy did not influence the neurologic response. The median time to neurologic progression was 7 months (range 2-15, months). RT for cerebral recurrence was required in 2 patients. The median survival of patients with brain metastases was 5.6 months (95% confidence interval, 4.4-6.8 months). Intracranial hemorrhagic complications were not observed. CONCLUSIONS: The current results indicated that it is feasible to treat patients who have advanced melanoma and small brain metastasis with temozolomide as the single treatment. The small subset of patients with systemic response usually showed durable stabilization or a response of brain metastasis. With this approach, neurologic disease can be controlled, and cranial irradiation may be deferred and even withheld in most of patients.     

Abdominal pain with a microvascular cause

A 50-year-old Caucasian man was admitted with unexplained abdominal pain. Based on CT scan results, a perforation resulting from colitis of the transverse colon was suspected. Additional colonoscopy showed ulcerative stenosis of the transverse colon. Initially, the patient recovered on conservative treatment; however, because of persistent stenosis, an elective resection of the transverse colon was performed. A CT scan revealed free gas around the anastomosis on the fourth postoperative day. Relaparotomy revealed leakage and diffuse ischaemic changes around the anastomosis as well as perforations at different sites on the small bowel. Postoperative CT angiography showed stenosis of the proximal celiac trunk, which was successfully stented. Digital subtraction angiography (DSA) revealed luminal irregularities of the superior mesenteric, the middle colic as well as the main hepatic and splenic arteries. Eventually, the clinical presentation and surgical, radiologic and histopathological findings together led to the diagnosis of polyarteritis nodosa (PAN). Aggressive treatment with glucocorticoids and cyclophosphamide resulted in the resolution of symptoms and the disappearance of radiologic abnormalities.     

Effectiveness of a short web-based film targeting parental oral health knowledge in a well-child care setting

Young children rely on their parents with respect to oral health routines. However, parental knowledge on this topic is often insufficient. Well-child care may be an excellent route to reach parents because almost all of them attend. To evaluate the effectiveness of an 8.5 min web-based film about oral health, provided by well-child care, a non-blinded quasi-experimental study was performed. Parents attending well-child care clinics in the Netherlands were assigned to an intervention (n = 88) or control group (n = 41). The control group received care as usual. We measured parental knowledge of oral health with a questionnaire (range of scores 1–12) before and directly after the intervention, and 6 months later, and assessed differences between the intervention and the control group. Parental oral health knowledge improved after watching the film: the intervention group’s mean score of 11.1 (SD 1.3) was greater than the mean score of 7.1 (SD 2.0) of the control group (Cohen's d = 2.64). Scores remained higher in the intervention group 6 months after watching the film (mean 9.1, SD 1.3) than before (Cohen's d = 1.25). A web-based educational film delivered in a well-child care setting can be an effective way to address oral health and to improve parental knowledge.     

Stretch reflex responses in Complex Regional Pain Syndrome-related dystonia are not characterized by hyperreflexia

ObjectiveTo evaluate if hyperreflexia (exaggerated reflexes) due to disinhibition is associated with dystonia in Complex Regional Pain Syndrome (CRPS).MethodsStretch reflexes at the wrist were assessed in healthy controls (n = 10) and CRPS-patients with dystonia (n = 10). Subjects exerted a wrist flexion torque of 5% of maximum voluntary contraction torque (T_MVC) to a manipulandum which applied ramp-and-hold stretches to the wrist flexors. Since reflex responses scale with background contraction, controls additionally performed the task at 1% and 3% T_MVC to attain similar torques as patients who have reduced T_MVC.The M1 onset and the magnitudes of the short latency M1 and long latency M2 were assessed using the electromyographic signals (EMG) of the flexor carpi radialis. EMG of the extensor carpi radialis was recorded to monitor cocontraction.ResultsCompared to controls, patients had a substantially reduced T_MVC. Ramp velocity had a significant effect on M1 onset time and magnitude.ConclusionsSince M1 magnitude decreased with flexion torque, no significant difference was found between patients and controls at 5% T_MVC, while comparison at similar absolute torques (controls at 1% T_MVC) resulted in significantly smaller M1 magnitudes for patients with dystonia.SignificanceThis study suggests that CRPS-patients with dystonia are not hyperreflexive.     

Alpha/beta ratio: A dose range dependence study

PURPOSE: To investigate the dependence of the alpha/beta ratio determined from in vitro survival curves on the dose ranges. METHODS: Detailed clonogenic cell survival experiments were used to determine the least squares estimators for the linear quadratic model for different dose ranges. The cell lines used were CHO AA8, a Chinese hamster fibroblast cell line; U-373 MG, a human glioblastoma cell line; and CP3 and DU-145, two human prostate carcinoma cell lines. The alpha, beta, and alpha/beta ratio behaviors, combined with a goodness-of-fit analysis and Monte Carlo simulation of the experiments, were assessed within different dose regions. RESULTS: Including data from the low-dose region has a significant influence on the determination of the alpha, beta, and alpha/beta ratio from in vitro survival curve data. In this region, the values are poorly determined and have significant variability. The mid-dose region is characterized by more precise and stable values and is in agreement with the linear quadratic model. The high-dose region shows relatively small statistical error in the fitted parameters but the goodness-of-fit and Monte Carlo analyses showed poor quality fits. CONCLUSION: The dependence of the fitted alpha and beta on the dose range has an impact on the alpha/beta ratio determined from the survival data. The low-dose region had a significant influence that could be a result of a strong linear, rather than quadratic, component, hypersensitivity, and adaptive responses. This dose dependence should be interpreted as a caution against using inadequate in vitro cell survival data for alpha/beta ratio determination.