CXCR4 chemokine receptors (CD184) and alpha4beta1 integrins mediate spontaneous migration of human CD34+ progenitors and acute myeloid leukaemia cells beneath marrow stromal cells (pseudoemperipolesis)

Marrow stromal cells play an important role in regulating the development and proliferation of haematopoietic stem cells (HSC) within the marrow microenvironment. However, the molecular mechanisms of stem cell-stromal cell interactions are not fully understood. We observed that mobilized peripheral blood and cord-blood-derived CD34+ progenitor cells, or CD34+ acute myeloid leukaemia (AML) cells spontaneously migrated beneath marrow stromal cells, an in vitro migration phenomenon termed pseudoemperipolesis. In contrast, the CD34+ myeloid leukaemia cell line, Kasumi-1, did not display pseudoemperipolesis. Cord blood CD34+ cells had a higher capacity than granulocyte-colony-stimulating-factor-mobilized CD34+ cells for pseudoemperipolesis (28.7 +/- 12%vs 18.1 +/- 6.1% of input cells within 24 h, mean +/- SD, n = 8), whereas 9.4 +/- 12.6% (mean +/- SD, n = 10) of input AML cells displayed this phenomenon. Pseudoemperipolesis of CD34+ progenitor and AML cells was significantly inhibited by pertussis toxin and antibodies to the CXCR4 chemokine receptor (CXCR4, CD184), but not control antibodies. Moreover, CD34+ and AML cell migration was significantly inhibited by a CS1 peptide that blocks alpha4beta1 integrin binding, but not by a control peptide, in which the fibronectin binding motif was scrambled. Pseudoemperipolesis was associated with an increased proliferation of migrated CD34+ progenitor cells but not AML cells within the stromal layer, demonstrated by cell cycle analysis and cell division tracking. We conclude that alpha4beta1 integrin binding and CXCR4 chemokine receptor activation are prerequisites for the migration of CD34+ haematopoietic progenitors and AML cells beneath marrow stromal cells. These observations suggest a central role of marrow stromal cells for HSC trafficking and homing within the marrow microenvironment.     

High-resolution x-ray structure of human aquaporin 5

Human aquaporin 5 (HsAQP5) facilitates the transport of water across plasma membranes and has been identified within cells of the stomach, duodenum, pancreas, airways, lungs, salivary glands, sweat glands, eyes, lacrimal glands, and the inner ear. AQP5, like AQP2, is subject to posttranslational regulation by phosphorylation, at which point it is trafficked between intracellular storage compartments and the plasma membrane. Details concerning the molecular mechanism of membrane trafficking are unknown. Here we report the x-ray structure of HsAQP5 to 2.0-Å resolution and highlight structural similarities and differences relative to other eukaryotic aquaporins. A lipid occludes the putative central pore, preventing the passage of gas or ions through the center of the tetramer. Multiple consensus phosphorylation sites are observed in the structure and their potential regulatory role is discussed. We postulate that a change in the conformation of the C terminus may arise from the phosphorylation of AQP5 and thereby signal trafficking.     

Is there a need for dietary measures to further reduce LDL cholesterol in patients with type II diabetes mellitus on statin therapy?

PURPOSE: To study the need for dietary measures to further reduce LDL cholesterol in patients with type 2 diabetes mellitus on statin therapy. METHODS: A Pubmed, Embase, CINAHL and Cochrane library search was performed to identify relevant articles. After critical appraisal six articles were ranked according to relevance, validation and level of evidence. RESULTS: There were no studies performed among type II diabetics. Among patients with hypercholesterolaemia, diet led to an additional LDL reduction from 0.20 to 0.88 mmol/l, translating into 23% reduction in vascular risk. CONCLUSION: We recommend a low-fat diet on top of statin therapy in patients with type 2 diabetes mellitus assuming that effects found in hypercholesterolaemic patients also apply to type 2 diabetics.     

Inhibition of WISE Preserves Renal Allograft Function

Wnt-modulator in surface ectoderm (WISE) is a secreted modulator of Wnt signaling expressed in the adult kidney. Activation of Wnt signaling has been observed in renal transplants developing interstitial fibrosis and tubular atrophy; however, whether WISE contributes to chronic changes is not well understood. Here, we found moderate to high expression of WISE mRNA in a rat model of renal transplantation and in kidneys from normal rats. Treatment with a neutralizing antibody against WISE improved proteinuria and graft function, which correlated with higher levels of β-catenin protein in kidney allografts. In addition, treatment with the anti-WISE antibody reduced infiltration of CD68⁺ macrophages and CD8⁺ T cells, attenuated glomerular and interstitial injury, and decreased biomarkers of renal injury. This treatment reduced expression of genes involved in immune responses and in fibrogenic pathways. In summary, WISE contributes to renal dysfunction by promoting tubular atrophy and interstitial fibrosis.The development of interstitial fibrosis and tubular atrophy with ongoing inflammation is a major risk for progressive graft dysfunction eventually leading to the failure of the majority of renal transplants. Although multiple approaches are available for preventing or at least blunting immune responses, there is currently no effective treatment for the development and progression of interstitial fibrosis and tubular atrophy.¹ The formation of matrix proteins in parallel to a progressive loss of graft function is a hallmark of chronic allograft dysfunction (CAD). Several pathways, including those involving TGF-β and angiotensin receptor, have been found to promote fibrosis and thus significant efforts have been made to evaluate the potential utility of these two pathway inhibitors for CAD. TGF-β has been shown to play an important role in epithelial-mesenchymal transition (EMT), which may, in turn, promote deteriorating structural changes characteristic for CAD.² However, inhibiting TGF-β may, because of its immunomodulatory effects, carry the risk of augmenting inflammation.³ Angiotensin II (AngII) is a growth factor that activates the Smad pathway during EMT involving TGF-β.⁴ AngII receptor antagonists were shown to reduce BP, proteinuria, and fibrosis in some studies.^5,6 However, the application of AngII receptor antagonists is known to be associated with intimal hyperplasia and deteriorating renal function, thus making its application in CAD challenging.⁷Wnt signaling is tightly regulated during kidney development and plays an important role in the formation of various structures of the developing kidney.^8–11 In normal adult kidneys, Wnt signaling is progressively downregulated once the developmental phase is completed.¹² Activation of Wnt signaling has been reported in a variety of human disease processes, including interstitial pulmonary fibrosis,¹³ and in transplanted kidneys undergoing interstitial fibrosis and tubular atrophy.¹⁴ To understand the contribution of Wnt-modulator in surface ectoderm (WISE) on tubular atrophy and interstitial fibrosis, we generated a potent rat inhibitory antibody to rat WISE, allowing long-term treatment while minimizing immune responses toward the injected antibody. Prophylactic treatment with a rat anti-WISE antibody, referred to hereafter as anti-WISE, reduced inflammatory infiltration, improved renal function, and reduced structural graft deterioration over a 6-month observation period. Serum biomarker and changes in gene expression suggested improvements in tubular epithelial integrity as well as decreases in profibrotic and inflammatory pathways, respectively. The improvement in graft function in our studies was associated with increased β-catenin levels. Moreover, WISE protein modulated Wnt signaling in vitro in a context-dependent manner, and directly affected E-cadherin expression and α-smooth muscle actin (α-SMA) expression in renal epithelial cells and interstitial fibroblasts.     

Using a Delphi process to develop an effective train-the-trainers program to train health and social care professionals throughout Europe

Research has shown that developing a Train-the-Trainers (TTT) program is important if agencies are to implement guidelines, but the most effective way to deliver a TTT program remains unanswered. This article presents data from a 3-round Internet-based Delphi process, which was used to help develop consensus-based guidelines for a TTT programme to deliver to health and social care professionals throughout Europe a curriculum on traumatic stress. In Round 1, 74 experts rated the importance of statements relating to the TTT field and then reassessed their scores in the light of others' responses in subsequent rounds. Forty-one (67%) of 61 statements achieved consensus (defined as having a mean score >7 or < 3 on the 0-9 rating scales used and 70% of participants scoring 7 and above or 3 and below) for inclusion. Key TTT components included interactive and practical presentations, delivery to groups of 7-12 people over 2 days, external and local expert facilitation, course manuals, refresher courses, and supervision. The Delphi process allowed a consensus to be achieved in an area in which there are limitations in the current evidence.     

Arteriolar vascular smooth muscle cell differentiation in benign nephrosclerosis

Background Benign nephrosclerosis (bN) is the most prevalent form of hypertensive damage in kidney biopsies. It is defined by early hyalinosis and later fibrosis of renal arterioles. Despite its high prevalence, very little is known about the contribution of arteriolar vascular smooth muscle cells (VSMCs) to bN. We examined classical and novel candidate markers of the normal contractile and the pro-fibrotic secretory phenotype of VSMCs in arterioles in bN. Methods Sixty-three renal tissue specimens with bN and eight control specimens were examined by immunohistochemistry for the contractile markers caldesmon, alpha-smooth muscle actin (alpha-SMA), JunB, smoothelin and the secretory marker S100A4 and by double stains for caldesmon or smoothelin with S100A4. Results Smoothelin immunostaining showed an inverse correlation with hyalinosis and fibrosis scores, while S100A4 correlated with fibrosis scores only. Neither caldesmon, alpha-SMA nor JunB correlated with hyalinosis or fibrosis scores. Cells in the arteriolar wall were exclusively positive either for caldesmon/smoothelin or S100A4. Conclusions This is the first systematic analysis of VSMC differentiation in bN. The results suggest that smoothelin is the most sensitive marker for the contractile phenotype and that S100A4 could be a novel marker for the secretory phenotype in vivo. The other markers did not seem to differentiate these phenotypes in bN. Thus, VSMC phenotype markers should be defined in the context of the vessel segment and disease under examination. S100A4 could not only be a marker of pro-fibrotic secretory VSMCs in bN but also an important mediator of arteriolar fibrosis.     

NBAS Variants Are Associated with Quantitative and Qualitative NK and B Cell Deficiency

Purpose

Biallelic pathogenic NBAS variants manifest as a multisystem disorder with heterogeneous clinical phenotypes such as recurrent acute liver failure, growth retardation, and susceptibility to infections. This study explores how NBAS-associated disease affects cells of the innate and adaptive immune system.

Methods

Clinical and laboratory parameters were combined with functional multi-parametric immunophenotyping methods in fifteen NBAS-deficient patients to discover possible alterations in their immune system.

Results

Our study revealed reduced absolute numbers of mature CD56^dim natural killer (NK) cells. Notably, the residual NK cell population in NBAS-deficient patients exerted a lower potential for activation and degranulation in response to K562 target cells, suggesting an NK cell–intrinsic role for NBAS in the release of cytotoxic granules. NBAS-deficient NK cell activation and degranulation was normalized upon pre-activation by IL-2 in vitro, suggesting that functional impairment was reversible. In addition, we observed a reduced number of naïve B cells in the peripheral blood associated with hypogammaglobulinemia.

Conclusion

In summary, we demonstrate that pathogenic biallelic variants in NBAS are associated with dysfunctional NK cells as well as impaired adaptive humoral immunity.