Disease-specific ex vivo stimulation of whole blood for cytokine production: applications in the study of tuberculosis

As a simple method for field studies to assess the cytokine-status of patients with tuberculosis (TB), the use of whole blood instead of isolated cells has advantages, especially since the risk of contamination is minimal. Therefore, cytokine production in whole blood cultures was determined using non-specific and disease-specific stimuli. Heparinized blood from healthy volunteers was either incubated in closed vacutainer tubes or in tissue culture wells after dilution in culture medium. Dose-response and kinetics were investigated for the production of TNFalpha, IL-1beta, IL-1ra, IL-10 and IFNgamma. Patients with TB and healthy individuals were examined for IFN-gamma production in whole blood. In the absence of a stimulus, the production of cytokines is negligible in whole blood cultures. LPS induces the production of TNFalpha, IL-1beta, IL-1ra and IL-10; PHA induces the production of IFNgamma and IL-10. Live BCG induces the production of proinflammatory cytokines, irrespective of tuberculin skin status. In contrast, PPD and MTB-culture filtrate induce production of IFNgamma in skin-test positive and not in skin-test negative healthy subjects. Five out of 13 patients with TB had a low antigen-specific IFNgamma production, suggestive of a minimal or absent specific T-cell response. For most purposes, cultures in closed vacutainer tubes are optimal. If one wishes to focus on T-cell cytokines or if only small volumes of blood are available, dilution of whole blood in culture medium before incubation in tissue culture wells may be preferable.     

Promoting wellness for women with multiple sclerosis.

There is increasing interest in wellness programs among health care providers and consumers. A unique intervention program was developed to promote wellness for women with multiple sclerosis (MS). Key processes include: provision of accurate knowledge that is specific to health promotion within the context of MS; enhancement of self-efficacy for health behaviors and individualized goal setting and monitoring. Participants have reported positive changes as a result of this two-phase (knowledge/skill building classes and telephone follow-up) intervention.     

Fas-FasL interactions modulate host defense against systemic Candida albicans infection.

Fas-FasL costimulation modulates the production of proinflammatory cytokines, and MRL/lpr mice, which lack a functional Fas molecule, produce more proinflammatory cytokines. This study found that Fas-FasL interactions are involved in host defense against lethal infection with Candida albicans. Macrophages of MRL/lpr mice produced significantly more tumor necrosis factor and interleukin-1 after stimulation with C. albicans than did control MRL+/+ macrophages. Mortality of Fas-deficient mice with disseminated candidiasis was significantly lower than control animals' because of decreased fungal load and inhibition of the formation of invasive hyphae in their organs. Increased recruitment of neutrophils at the infection site appeared to be responsible for these effects. In contrast, phagocytosis and killing of C. albicans by neutrophils of MRL/lpr and MRL+/+ mice was similar. Absence of Fas-FasL interactions leads to increased cytokine production after C. albicans stimulation, protecting mice against disseminated candidiasis.     

Comparison of serum and whole blood levels of cytomegalovirus and Epstein–Barr virus DNA

Abstract: The monitoring of viral DNA levels after transplantation is crucial for prevention of complications from cytomegalovirus (CMV) or Epstein–Barr virus (EBV) infection but there is no consensus as to which matrix is the most adequate. To compare serum and whole blood (WB) as specimens for measuring viral DNA, clinical samples from a 3‐year period were studied, with focus on cases where serum and WB were drawn on the same day. In 1896 paired serum and WB samples, CMV DNA was detected in both specimen types in 472 samples with 0.18 log higher levels (P<0.001) in WB than in serum (median level 2.73 vs. 2.56 log copies/mL), and in only either serum or WB in 127 and 108 samples, respectively, generally at levels below 1000 copies/mL. In 664 paired samples, EBV DNA was detected in both serum and WB in 160 samples, with 1.48 log higher levels (P<0.001) in WB (median 4.2 vs. 2.4 log copies/mL), in only WB in 227 cases with a median at 3.0 log copies/mL, and only in serum in 14 samples at low levels. The correlation between serum and WB DNA levels was weaker for EBV than for CMV (R² 0.31 vs. 0.74). We conclude that either serum or WB may be used for monitoring CMV and EBV DNA levels, that EBV DNA is detected post transplant in >50% of WB samples and at 30 times higher levels than in serum, and that post‐transplantation lymphoproliferative disorder (PTLD) may develop without further increase of EBV DNA in WB. Identification of PTLD may require EBV DNA testing in both specimen types or complementary tests.     

Landmark‐based software for anatomical measurements: A precision study

The aim of this study was to develop a software program, called Landmarker, which would aid studies of complex anatomical morphometry by simplifying the manual identification of landmarks in 3D images. We also tested its precision on routine magnetic resonance imaging (MRI) scans. To understand human biological variation, there is a need to identify morphological characteristics from the exterior and the interior of human anatomy. MRI, as opposed to other radiographic methods (mainly based on X‐ray techniques), supplies good soft tissue contrast, which allows for more complex assessments than what bony landmarks can provide. Because automation of this assessment is highly demanding, one of the primary goals for the new software was to enable more rapid identification of landmark sets in 3D image data. Repeat acquisition of head MRIs having a resolution of 0.94 × 0.94 × 1.20 mm³ were performed on 10 volunteers. Intra‐ and interoperator, as well as interacquisition variations of manual identification of exterior, craniofacial interior, and brain landmarks were studied. The average distances between landmarks were <1.8 mm, <2.3 mm, and <2.0 mm in the intra‐ and interoperator, and interacquisition evaluations, respectively. This study presents new software for time efficient identification of complex craniofacial landmarks in 3D MRI. To the best of our knowledge, no evaluation of software for rapid landmark‐based analysis of complex anatomies from 3D MR data has yet been presented. This software may also be useful for studies in other anatomical regions and for other types of image data. Clin. Anat. 22:456–462, 2009. © 2009 Wiley‐Liss, Inc.     

Severe osteomyelitis due to the zygomycete Apophysomyces elegans.

We describe a previously healthy 69-year-old man presenting with osteomyelitis of the humerus due to the zygomycete Apophysomyces elegans. The infection was acquired in Aruba, The Netherlands Antilles. The skin provided the most likely portal of entry, although there was no history of a traumatic inoculation. The patient had no history of diabetes, and no underlying immune defects were found. Despite treatment with 7.9 g of amphotericin B, an interthoracoscapular amputation proved necessary to curtail the rapid spread of the fungus in this immunocompetent host.     

Psoriasis under the microscope

Histopathology is a major diagnostic tool in dermatology, particularly in psoriasiform diseases. Morphological studies showed that the initial event in psoriatic lesions is perivascular infiltrate, followed by dilatation of superficial papillary vessels. Proliferation of keratinocytes and neutrophil exocytosis are secondary events. Fully developed psoriasis has a very characteristic pattern, which includes elongation of rete ridges leading to regular acanthosis, oedema of the papillary dermis associated with tortuous dilated vessels, thinning of suprapapillar area, decreased thickness of granular layer, and exocytosis of neutrophils in the spinous layer (Kogoj's pustule) or in the cornified parakeratotic layer (Munro microabscesses). Pustular psoriasis is characterized by large or confluent intra‐epidermal multilocular pustules. Whatever the clinical variant of psoriasis, common morphological signs suggest that it is basically a unique pathological process, with many possible presentations according to various factors such as age, size and localization of lesions, or therapy. Similar microscopic elementary lesions indicate that Hallopeau's acrodermatitis continua, Reiter's disease and geographical tongue are variants of psoriasis. Because of the many faces of the disease, psoriasis can resemble many other squamous or pustular disorders. Differential diagnosis by microscopic analysis is based on pattern analysis, PAS (Periodic Acid Schiff) staining to rule out fungal infection, and immunohistochemistry to characterize lymphocytic infiltrate. Psoriasis is one of the most common inflammatory skin diseases. In its characteristic presentation, psoriasis comprises well‐circumscribed red scaly papules and plaques. In this form, the disease is generally easy to identify, especially when the elbows, knees and scalp are affected. Nevertheless, the term ‘psoriasis’ includes more clinical variants than any other inflammatory dermatosis: psoriasis vulgaris vs. pustular, localized vs. generalized, topographic variants, mucous membranes involvement, hair and nail lesions. Although some of these conditions might be extremely different from psoriasis vulgaris, common pathological findings can be identified in all of them. Microscopic analysis of psoriatic lesions may therefore help clinicians to make the diagnosis and to understand that, whatever the clinical presentation, signs and symptoms are mainly due to a unique pathological process.     

Invasive zygomycosis: notably in diabetes mellitus and iron overload

The incidence of invasive zygomycosis, a severe and often life-threatening infection, is increasing. The most common manifestations are pulmonary infection (following anti-cancer chemotherapy or haematopoietic stem-cell transplant) and invasive rhinocerebral infection (in patients with diabetes mellitus or iron overload). Iron metabolism plays an important role in the pathogenesis of infection in these high-risk populations. Rapid diagnosis, reversal of the underlying predisposition and timely surgical debridement are the underlying principles of therapy for this disease.     

CD4+ T Cell–mediated Granulomatous Pathology in Schistosomiasis Is Downregulated by a B Cell–dependent Mechanism Requiring Fc Receptor Signaling

The effector functions of CD4⁺ T lymphocytes are generally thought to be controlled by distinct populations of regulatory T cells and their soluble products. The role of B cells in the regulation of CD4-dependent host responses is less well understood. Hepatic egg granuloma formation and fibrosis in murine schistosomiasis are dependent on CD4⁺ lymphocytes, and previous studies have implicated CD8⁺ T cells or cross-regulatory cytokines produced by T helper (Th) lymphocytes as controlling elements of this pathologic process. In this report, we demonstrate that B cell–deficient (μMT) mice exposed to Schistosoma mansoni develop augmented tissue pathology and, more importantly, fail to undergo the spontaneous downmodulation in disease normally observed during late stages of infection. Unexpectedly, B cell deficiency did not significantly alter T cell proliferative response or cause a shift in the Th1/Th2 balance. Since schistosome-infected Fc receptor–deficient (FcR γ chain knockout) mice display the same exacerbated egg pathology as that observed in infected μMT mice, the B cell– dependent regulatory mechanism revealed by these experiments appears to require receptor-mediated cell triggering. Together, the data demonstrate that humoral immune response/FcR interactions can play a major role in negatively controlling inflammatory disease induced by CD4⁺ T cells.     

添加谷氨酰胺的全胃肠外营养对造血干细胞移植术后常见并发症的防治作用

目的:严重的黏膜损伤是诱发造血干细胞移植后出现并发症的一常见原因,已有证据显示谷氨酰胺能降低接受化疗患儿黏膜炎的发生率。观察谷氨酰胺对异基因外周造血干细胞移植患者并发症及恢复的影响。方法:选择于2002-03/2006-11在河南省血液病研究所接受同胞异基因外周造血干细胞移植的48例血液系统肿瘤患者。所有患者及其家属对治疗和实验均知情同意,并经医院伦理委员会批准。所有患者移植前均处于完全缓解状态,营养中等或良好,心、肝、肾功能正常,将48例患者随机分为标准化全胃肠外营养液组(标准组,n=13)和加用谷氨酰胺的全胃肠外营养液组(谷氨酰胺组,n=35)。待患者中性粒细胞升至1.0×109L-1,且无任何感染指征,进行异基因外周造血干细胞移植。造血干细胞输注后第1天开始给予全胃肠外营养与胃肠外营养联合谷氨酰胺双肽,至中性粒细胞≥1.0×109L-1,且无消化道症状时停用。观察两组患者中性粒细胞恢复时间、出层流室时间以及关于感染、急性移植物抗宿主病等情况有无差异。结果:48例患者均进入结果分析。两组患者营养物质的摄入基本相同,谷氨酰胺组有6例发生黏膜炎,标准组有11例,差异显著(P<0.05);谷氨酰胺组有1例发生严重腹泻,标准组有5例,差异显著(P<0.05);谷氨酰胺组有3例发生临床感染,标准组有7例,差异显著(P<0.05);标准组中性粒细胞≥0.5×109L-1的持续时间短于谷氨酰胺组(P>0.05);谷氨酰胺组抗生素治疗时间及无菌病房居住时间较标准组短(P<0.05);两组急性移植物抗宿主病发生率差异无统计学意义(P>0.05)。结论:添加谷氨酰胺的全胃肠外营养可改善异基因造血干细胞移植患者的营养状态,减少感染及肠损害,减少急性移植物抗宿主病的发生,有利于异基因移植患者恢复。