Induction of apoptosis of human primary osteoclasts treated with extracts from the medicinal plant <Emphasis Type="Italic">Emblica officinalis</Emphasis>

Background  

Osteoclasts (OCs) are involved in rheumatoid arthritis and in several pathologies associated with bone loss. Recent results support the concept that some medicinal plants and derived natural products are of great interest for developing therapeutic strategies against bone disorders, including rheumatoid arthritis and osteoporosis. In this study we determined whether extracts of Emblica officinalis fruits display activity of possible interest for the treatment of rheumatoid arthritis and osteoporosis by activating programmed cell death of human primary osteoclasts.     

Conformational analysis of heparin binding peptides

A properly engineered biomaterial for dental/orthopaedic applications must induce specific responses from the osteoblasts at the implant site. A most desirable response is an efficient adhesion, as it represents the first phase in the cell/material interaction and the quality of this phase will influence the cell's capacity to organize into a new functional tissue. The four osteoblast-adhesive peptides discussed in this paper are mapped on the 339-364 sequence (339MAPRPSLAKKQRFRHRNRKGYRSQRG364) located in the primary heparin-binding site of human vitronectin (HVP). Adsorbed on a polystyrene scaffold, these peptides display different adhesive activities towards osteoblasts. In this paper we report on the structural analysis in solution of the peptides through NMR and computational techniques. We find that the peptides with the highest adhesive activities display a hydrophobic patch opposite to the charged surface candidate to interact with heparin. These findings suggest that the peptides might adsorb on the polystyrene support in a favourable orientation for their activity. Furthermore, molecular models obtained for the four peptides in solution were used in rigid docking simulations with a heparin model. Assuming that the peptide solution conformations are not very different from the polystyrene-adsorbed structures, the simulations reveal that peptide adhesive activity is also affected by the number of ionic interactions and spacing between charged residues.     

Ligand and cytokine dependence of the immunosuppressive pathway of tryptophan catabolism in plasmacytoid dendritic cells

Murine plasmacytoid dendritic cells (pDCs) have been credited with a unique ability to express indoleamine 2,3-dioxygenase (IDO) function and mediate immunosuppression in specific settings; yet, the conditions of spontaneous versus induced activity have remained unclear. We have used maneuvers known to up-regulate IDO in different cell types and have examined the relative efficacy and mechanisms of the induced activity in splenic pDCs, namely, after specific receptor engagement by CTLA-4-Ig, CD200-Ig or CD28-Ig, the latter in combination with silenced expression of the suppressor of cytokine signaling 3 (SOCS3) gene. We found that pDCs (CD11c+ mPDCA-1+ 120G8+) do not express IDO and are not tolerogenic under basal conditions. B7-1 engagement by CTLA-4-Ig, CD200R1 engagement by CD200-Ig and B7-1/B7-2 engagement by CD28-Ig in SOCS3-deficient pDCs were each capable of initiating IDO-dependent tolerance via different mechanisms. IFN-gamma was the major cytokine responsible for CTLA-4-Ig effects, and type I IFNs for those of CD200-Ig. Immunosuppression by CD28-Ig in the absence of SOCS3 required IFN-gamma induction and IFN-like actions of IL-6. Therefore, although pDCs do not mediate IDO-dependent tolerance constitutively, multiple ligands and cytokines will contribute to the expression of a tolerogenic phenotype by pDCs in the mouse.     

Expression of interleukin-8 gene in inflammatory bowel disease is related to the histological grade of active inflammation.

Interleukin-8 (IL-8) is a potent cytokine for recruitment and activation of neutrophils. To visualize its distribution in the intestinal mucosa and to understand better its possible role in the induction and promotion of inflammatory bowel disease, expression of the IL-8 gene was analyzed in resected bowel segments of 14 patients with active Crohn's disease or ulcerative colitis. In situ hybridization with IL-8 anti-sense RNA probes revealed strong and specific signals in the histologically affected mucosa. The number of cells expressing IL-8 gene correlated with the histological grade of active inflammation. In accordance with the characteristic histological signs of active disease, IL-8-expressing cells were diffusely distributed over the entire affected mucosa in patients with ulcerative colitis, whereas in patients with Crohn's disease, IL-8-expressing cells showed a focal distribution pattern. Cells expressing IL-8 were mainly located at the base of ulcers, in inflammatory exudates on mucosal surfaces, in crypt abscesses, and at the border of fistulae. Analysis of semi-serial sections pointed to macrophages, neutrophils, and epithelial cells as possible sources of this cytokine in active inflammatory bowel disease. We consistently failed to detect IL-8 messenger RNA in the mucosa of uninvolved bowel segments and in normal-appearing control mucosa of patients with colon cancer. In contrast, tissue specimens from two patients with acute appendicitis displayed IL-8-expressing cells in the mucosa. These results support the notion that IL-8 plays and important but nonspecific role in the pathogenesis of inflammatory bowel disease and that the production of IL-8 messenger RNA is restricted to areas with histological signs of inflammatory activity and mucosal destruction.     

Lymphocyte Modulation in a Baboon Model of Immunosenescence

The age-related modulation of lymphocyte number and function was assessed in a nonhuman primate model consisting of healthy olive baboons (Papio cynocephalus anubis) of ages encompassing the entire life span of this species. The objectives of this study were to characterize an animal model of immunosenescence and to assess whether or not age should be considered when designing studies for the evaluation of vaccine candidates in baboons. Specifically the following parameters were assessed in baboons from 6 months to 26 years of age: relative numbers of B lymphocytes, CD4⁺ and CD8⁺ T lymphocytes, and T lymphocytes expressing CD28, CD25, and phytohemagglutinin-stimulated lymphoproliferative activity; and concentrations of total immunoglobulin, soluble interleukin-2 receptor α, and soluble CD30 in serum. There was a statistically significant effect of age on lymphocyte numbers. As age increased, relative B-cell numbers (ranging from 6 to 50%) decreased (P < 0.001) and relative T-cell numbers (ranging from 28 to 80%) increased (P < 0.001). The increase in T-cell numbers involved both the CD4⁺ and CD8⁺ subsets. In addition, there was a significant negative correlation of age with levels of soluble interleukin-2 receptor α in serum. Modulation of lymphocyte numbers appears to occur gradually during the entire baboon life span, thus suggesting the presence of an age-related developmentally regulated process. These findings indicate that baboons represent a potentially useful model to study selected phenomena related to immunosenescence. These findings also indicate that, when using the baboon model for vaccine or other experimental protocols requiring the assessment of immune responses, it would be appropriate to take into account the age of the animals in the study design.     

ALK Expression Defines a Distinct Group of T/Null Lymphomas (“ALK Lymphomas”) with a Wide Morphological Spectrum

The t(2;5)(p23;q35) translocation associated with CD30-positive anaplastic large cell lymphoma results in the production of a NPM-ALK chimeric protein, consisting of the N-terminal portion of the NPM protein joined to the entire cytoplasmic domain of the neural receptor tyrosine kinase ALK. The ALK gene products were identified in paraffin sections by using a new anti-ALK (cytoplasmic portion) monoclonal antibody (ALKc) that tends to react more strongly than a previously described ALK1 antibody with the nuclei of ALK-expressing tumor cells after microwave heating in 1 mmol/L ethylenediaminetetraacetic acid buffer, pH 8.0. The ALKc monoclonal antibody reacted selectively with 60% of anaplastic large cell lymphoma cases (60 of 100), which occurred mainly in the first three decades of life and consistently displayed a T/null phenotype. This group of ALK-positive tumors showed a wide morphological spectrum including cases with features of anaplastic large cell lymphoma “common” type (75%), “lymphohistiocytic” (10%), “small cell” (8.3%), “giant cell” (3.3%), and “Hodgkin’s like” (3.3%). CD30-positive large anaplastic cells expressing the ALK protein both in the cytoplasm and nucleus represented the dominant tumor population in the common, Hodgkin’s-like and giant cell types, but they were present at a smaller percentage (often with a perivascular distribution) also in cases with lymphohistiocytic and small cell features. In this study, the ALKc antibody also allowed us to identify small neoplastic cells (usually CD30 negative) with nucleus-restricted ALK positivity that were, by definition, more evident in the small cell variant but were also found in cases with lymphohistiocytic, common, and “Hodgkin’s-like” features. These findings, which have not been previously emphasized, strongly suggest that the neoplastic lesion (the NPM-ALK gene) must be present both in the large anaplastic and small tumor cells, and that ALK-positive lymphomas lie on a spectrum, their position being defined by the ratio of small to large neoplastic cells. Notably, about 15% of all ALK-positive lymphomas (usually of the common or giant cell variant) showed a cytoplasm-restricted ALK positivity, which suggests that the ALK gene may have fused with a partner(s) other than NPM. From a diagnostic point of view, detection of the ALK protein was useful in distinguishing anaplastic large cell lymphoma cases of lymphohistiocytic and small cell variants from reactive conditions and other peripheral T-cell lymphoma subtypes, as well as for detecting a small number of tumor cells in lymphohemopoietic tissues. In conclusion, ALK positivity appears to define a clinicopathological entity with a T/null phenotype (“ALK lymphomas”), but one that shows a wider spectrum of morphological patterns than has been appreciated in the past.     

Stress-activated protein kinases mediate cell migration in human airway epithelial cells

Airway epithelial cell (AEC) repair immediately after injury requires coordinated cell spreading and migration at the site of injury. Stress-activated protein kinases such as p38 MAPK and c-Jun N-terminal Protein Kinase (JNK) modulate several responses to cell stress and injury, but their role in AEC migration is not clear. We examined migration in confluent 16HBE14o(-) human AEC lines and in primary AEC grown on collagen-IV. Wounds were created by mechanical abrasion and followed to closure using digital microscopy. Inhibitors of either p38 extracellular signal-regulated kinase (ERK)1/2 (PD98059), mitogen-activated protein kinase (MAPK) (SB203580), or JNK (SP600125) could block cell migration substantially. Inhibiting JNK but not p38 MAPK or ERK1/2 blocked extension of cells into the wound region from the original line of injury. Initial migration was associated with phosphorylation of ERK, p38 MAPK, and JNK within 5-15 min. The downstream effector of p38, heat shock protein 27, also was phosphorylated rapidly after injury; phosphorylation could be blocked by prior treatment with SB203580 but not SP600125. The downstream effector of JNK, c-Jun, likewise was phosphorylated rapidly after injury and could be blocked by inhibiting JNK. Our data demonstrate that p38 MAPK, JNK, and ERK1/2 participate in the early stages of AEC migration.     

PG-M1: A New Monoclonal Antibody Directed against a Fixative-Resistant Epitope on the Macrophage-Restricted Form of the CD68 Molecule

A new anti-macrophage monoclonal antibody (PG-M1) was produced by immunizing BALB/c mice with fresh spleen cells from a patient with Gaucher's disease. PG-M1 reacts strongly with a fixative-resistant epitope of an intracytoplasmic molecule, selectively expressed by virtually all macrophages of the human body. Although attempts to immunoprecipitate the molecule recognized by PG-M1 have failed so far, the reactivity of the antibody with COS-1 and WOP cells transfected with a human complementary DNA clone encoding for the CD68 antigen suggests that PG-M1 is a new member of the CD68 cluster. However, unlike other CD68 antibodies (KP1, EBM11, etc.), which react with both macrophages and myeloid cells, PG-M1 detects a fixative-resistant epitope on the macrophage-restricted form of the CD68 antigen. In 957 routinely fixed, paraffin-embedded samples, PG-M1 showed a more restricted reactivity with elements of the monocyte/macrophage lineage than the previously described monoclonal antibodies MAC-387 (anti-calgranulins), KP1 (CD68) and Ki-M1P. Among hematological malignancies, PG-M1 only labels acute leukemias of M4 and M5 type and rare examples of malignant histiocytosis/true histiocytic sarcoma. In contrast, acute leukemias of the M1, M2, M3, M6, M7, and L1-L3 types, non-Hodgkin's lymphomas, and Hodgkin and Reed-Sternberg cells of Hodgkin's disease are consistently PG-M1-negative. In the daily diagnostic practice, PG-M1 seems to be particularly valuable for the diagnosis of myelomonocytic or monocytic leukemia and neoplasms of true histiocytic origin in routine paraffin sections.     

Diagnostic performance of aPS/PT antibodies in neuropsychiatric lupus and cardiovascular complications of systemic lupus erythematosus

Abstract

Background: Systemic lupus erythematosus (SLE) is associated with a constellation of complications affecting multiple organs, including neuropsychiatric manifestations (NPSLE) and ischaemic events, leading to increased long-term morbidity. Antiphospholipid antibodies (aPL) are a major determinant of vascular inflammation and thromboembolic risk. The diagnostic role of anti-phosphatidylserine/prothrombin (aPS/PT) antibodies in this setting is incompletely defined.

Aim: To verify whether aPS/PT add to diagnostics and disease stratification in patients with SLE with or without other aPL.

Methods: 131 consecutive patients were studied, including 20 patients with SLE and secondary antiphospholipid syndrome (APS). aPS/PT IgG and IgM were assessed through ELISA and patients were stratified based on the presence of other aPL, on their clinical and laboratory features at time of blood sampling and on their clinical history. Synthetic indices of disease activity, chronic damage and cardiovascular risk were calculated at time of venipuncture.

Results: Fifty-one (38.9%) patients with SLE had aPS/PT and 15 (11.5%) patients had aPS/PT as the only aPL (aPS/PT-only). aPS/PT-only patients had a significantly higher prevalence of NPSLE than quadruple aPL-negative patients (p?=?.007). Patients with aPS/PT were more likely to have a history of ischaemia, thrombocytopenia and Libman–Sacks’ endocarditis. The presence of aPS/PT also associated with previous accrual of at least one damage item (p?=?.043), but had limited predictive values for damage progression in the short term.

Conclusion: aPS/PT antibodies provide non-redundant information that could contribute to risk assessment and stratification of patients with SLE.