AKT signaling is important for proliferation and survival of tumor cells. The clinical significance of AKT activation in diffuse large B-cell lymphoma (DLBCL) is not well analyzed. Here, we assessed ...expression of phosphorylated AKT (p-AKT) in 522 DLBCL patients. We found high levels of p-AKT nuclear expression, observed in 24.3% of the study cohort, were associated with significantly worse progression-free survival and Myc and Bcl-2 overexpression. However, multivariate analysis indicated that AKT hyperactivation was not an independent factor. miRNA profiling analysis demonstrated that 63 miRNAs directly or indirectly related to the phosphatidylinositol 3-kinase/AKT/mechanistic target of rapamycin pathway were differentially expressed between DLBCLs with high and low p-AKT nuclear expression. We further targeted the AKT signaling using a highly selective AKT inhibitor MK-2206 in 26 representative DLBCL cell lines and delineated signaling alterations using a reverse-phase protein array. MK-2206 treatment inhibited lymphoma cell viability, and MK-2206 sensitivity correlated with AKT activation status in DLBCL cells. On MK-2206 treatment, p-AKT levels and downstream targets of AKT signaling were significantly decreased, however, likely because of the decreased feedback repression; Rictor and phosphatidylinositol 3-kinase expression and other compensatory pathways were also induced. This study demonstrates the clinical and therapeutic implication values of AKT hyperactivation in DLBCL and suggests that AKT inhibitors need to be combined with other targeted agents for DLBCL to achieve optimal clinical efficacy.
The relationships between the molecular abnormalities in mitochondrial respiratory chain complexes and their negative contributions to mitochondrial and cellular functions have been proved to be ...essential for better understandings in mitochondrial medicine. Herein, we established the method to identify disease phenotypic differences among patients with muscle histopathological cytochrome c oxidase (COX) deficiency, as one of the representative clinical features in mitochondrial diseases, by using patients' myoblasts that are derived from biopsied skeletal muscle tissues. We identified two obviously different severities in molecular diagnostic criteria of COX deficiency among patients: structurally stable, but functionally mild/moderate defect and severe functional defect with the disrupted COX holoenzyme structure. COX holoenzyme disorganization actually triggered several mitochondrial dysfunctions, including the decreased ATP level, the increased oxidative stress level, and the damaged membrane potential level, all of which lead to the deteriorated cellular growth, the accelerated cellular senescence, and the induced apoptotic cell death. Our cell-based in vitro diagnostic approaches would be widely applicable to understanding patient-specific pathomechanism in various types of mitochondrial diseases, including other respiratory chain complex deficiencies and other mitochondrial metabolic enzyme deficiencies.
Academic pathology departments will be dramatically affected by proposed United States federal government regulatory initiatives. Pathology research will be substantially altered if proposed changes ...to the Common Rule (Code of Federal Regulations: Protection of Human Subjects title 45 CFR 46) and regulations governing the return of individual research results are approved and finalized, even more so now that the Precision Medicine initiative has been launched. Together, these changes are disruptive influences on academic pathology research as we know it, straining limited resources and compromising advances in diagnostic and academic pathology. Academic research pathologists will be challenged over the coming years and must demonstrate leadership to ensure the continued availability of and the ethical use of research pathology specimens.
Activated T cells play crucial roles in the pathogenesis of autoimmune diseases, including lupus nephritis (LN). The activation of calcineurin/nuclear factor of activated T cells (NFAT) and STAT4 ...signaling is essential for T cells to perform various effector functions. Here, we identified the growth factor midkine (MK; gene name, Mdk ) as a novel regulator in the pathogenesis of 2,6,10,14-tetramethylpentadecane-induced LN via activation of NFAT and IL-12/STAT4 signaling. Wild-type ( Mdk+/+ ) mice showed more severe glomerular injury than MK-deficient ( Mdk−/− ) mice, as demonstrated by mesangial hypercellularity and matrix expansion, and glomerular capillary loops with immune-complex deposition. Compared with Mdk−/− mice, the frequency of splenic CD69+ T cells and T helper (Th) 1 cells, but not of regulatory T cells, was augmented in Mdk+/+ mice in proportion to LN disease activity, and was accompanied by skewed cytokine production. MK expression was also enhanced in activated CD4+ T cells in vivo and in vitro . MK induced activated CD4+ T cells expressing CD69 through nuclear activation of NFAT transcription and selectively increased in vitro differentiation of naive CD4+ T cells into Th1 cells by promoting IL-12/STAT4 signaling. These results suggest that MK serves an indispensable role in the NFAT-regulated activation of CD4+ T cells and Th1 cell differentiation, eventually leading to the exacerbation of LN.
The clinical progression of Alzheimer disease (AD) is associated with the accumulation of tau neurofibrillary tangles, which may spread throughout the cortex by interneuronal tau transfer. If so, ...targeting extracellular tau species may slow the spreading of tau pathology and possibly cognitive decline. To identify suitable target epitopes, we tested the effects of a panel of tau antibodies on neuronal uptake and aggregation in vitro . Immunodepletion was performed on brain extract from tau-transgenic mice and postmortem AD brain and added to a sensitive fluorescence resonance energy transfer–based tau uptake assay to assess blocking efficacy. The antibodies reduced tau uptake in an epitope-dependent manner: N-terminal (Tau13) and middomain (6C5 and HT7) antibodies successfully prevented uptake of tau species, whereas the distal C-terminal–specific antibody (Tau46) had little effect. Phosphorylation-dependent (40E8 and p396) and C-terminal half (4E4) tau antibodies also reduced tau uptake despite removing less total tau by immunodepletion, suggesting specific interactions with species involved in uptake. Among the seven antibodies evaluated, 6C5 most efficiently blocked uptake and subsequent aggregation. More important, 6C5 also blocked neuron-to-neuron spreading of tau in a unique three-chamber microfluidic device. Furthermore, 6C5 slowed down the progression of tau aggregation even after uptake had begun. Our results imply that not all antibodies/epitopes are equally robust in terms of blocking tau uptake of human AD-derived tau species.
The blood–brain barrier (BBB) regulates cerebrovascular permeability and leakage of blood‐derived fibrinogen. Dysfunction of the BBB has been associated with cerebral arteriolosclerosis small vessel ...disease (SVD) and white matter lesions (WML). Furthermore, BBB dysfunction is associated with the pathogenesis of Alzheimer’s disease (AD) with the presence of CSF plasma proteins suggested to be a potential biomarker of AD. We aimed to determine if extravascular fibrinogen in the white matter was associated with the development of AD hallmark pathologies, i.e., hyperphosphorylated tau (HPτ) and amyloid‐β (Aβ), as well as SVD, cerebral amyloid angiopathy (CAA) and measures of white matter damage. Using human post‐mortem brains, parietal tissue from 20 AD and 22 non‐demented controls was quantitatively assessed for HPτ, Aβ, white matter damage severity, axonal density, demyelination and the burden of extravascular fibrinogen in both WML and normal appearing white matter (NAWM). SVD severity was determined by calculating sclerotic indices. WML‐ and NAWM fibrinogen burden was not significantly different between AD and controls nor was it associated with the burden of HPτ or Aβ pathology, or any measures of white matter damage. Increasing severity of SVD was associated with and a predictor of both higher WML‐ and NAWM fibrinogen burden (all P < 0.05) in controls only. In cases with minimal SVD NAWM fibrinogen burden was significantly higher in the AD cases (P < 0.05). BBB dysfunction was present in both non‐demented and AD brains and was not associated with the burden of AD‐associated cortical pathologies. BBB dysfunction was strongly associated with SVD but only in the non‐demented controls. In cases with minimal SVD, BBB dysfunction was significantly worse in AD cases possibly indicating the influence of CAA. In conclusion, extravascular fibrinogen is not associated with AD hallmark pathologies but indicates SVD, suggesting that the presence of fibrinogen in the CSF is not a surrogate marker for AD pathology.
The progressive loss of pancreatic β‐cell mass that occurs in both type 1 and type 2 diabetes is a primary factor driving efforts to identify strategies for effectively increasing, enhancing or ...restoring β‐cell mass. While factors that seem to influence β‐cell proliferation in specific contexts have been described, reliable stimulation of human β‐cell proliferation has remained a challenge. Importantly, β‐cells exist in the context of a complex, integrated pancreatic islet microenvironment where they interact with other endocrine cells, vascular endothelial cells, extracellular matrix, neuronal projections and islet macrophages. This review highlights different components of the pancreatic microenvironment, and reviews what is known about how signaling that occurs between β‐cells and these other components influences β‐cell proliferation. Future efforts to further define the role of the pancreatic islet microenvironment on β‐cell proliferation may lead to the development of successful approaches to increase or restore β‐cell mass in diabetes.
The colonic epithelium facilitates host-microorganism interactions to control mucosal immunity, coordinate nutrient recycling and form a mucus barrier. Breakdown of the epithelial barrier underpins ...inflammatory bowel disease (IBD). However, the specific contributions of each epithelial-cell subtype to this process are unknown. Here we profile single colonic epithelial cells from patients with IBD and unaffected controls. We identify previously unknown cellular subtypes, including gradients of progenitor cells, colonocytes and goblet cells within intestinal crypts. At the top of the crypts, we find a previously unknown absorptive cell, expressing the proton channel OTOP2 and the satiety peptide uroguanylin, that senses pH and is dysregulated in inflammation and cancer. In IBD, we observe a positional remodelling of goblet cells that coincides with downregulation of WFDC2-an antiprotease molecule that we find to be expressed by goblet cells and that inhibits bacterial growth. In vivo, WFDC2 preserves the integrity of tight junctions between epithelial cells and prevents invasion by commensal bacteria and mucosal inflammation. We delineate markers and transcriptional states, identify a colonic epithelial cell and uncover fundamental determinants of barrier breakdown in IBD.
It has been recognized that molecular classifications will form the basis for neuropathological diagnostic work in the future. Consequently, in order to reach a diagnosis of Alzheimer's disease (AD), ...the presence of hyperphosphorylated tau (HP‐tau) and β‐amyloid protein in brain tissue must be unequivocal. In addition, the stepwise progression of pathology needs to be assessed. This paper deals exclusively with the regional assessment of AD‐related HP‐tau pathology. The objective was to provide straightforward instructions to aid in the assessment of AD‐related immunohistochemically (IHC) detected HP‐tau pathology and to test the concordance of assessments made by 25 independent evaluators. The assessment of progression in 7‐µm‐thick sections was based on assessment of IHC labeled HP‐tau immunoreactive neuropil threads (NTs). Our results indicate that good agreement can be reached when the lesions are substantial, i.e., the lesions have reached isocortical structures (stage V–VI absolute agreement 91%), whereas when only mild subtle lesions were present the agreement was poorer (I–II absolute agreement 50%). Thus, in a research setting when the extent of lesions is mild, it is strongly recommended that the assessment of lesions should be carried out by at least two independent observers.
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