[Determination of the healing effect of Piper aduncum (spiked pepper or matico) on human fibroblasts]. / Determinación del efecto cicatrizante de Piper aduncum (matico) en fibroblastos humanos.

OBJECTIVES.: To evaluate the healing effect of a Piper aduncum ethanol-water extract on an adult human dermal fibroblast cell line (hDFa). MATERIALS AND METHODS.: After obtaining the extract via solid-liquid extraction, concentration, and lyophilization, extract proteins were purified using reverse phase high-performance liquid chromatography, identified using tandem mass spectrometry of tryptic peptides, and analyzed using MALDI-TOF-TOF on an ABSciex4800 mass spectrometer. Half maximum effective concentration values (EC50), half maximum inhibiting concentration (IC50), and percentages of cell proliferation were determined using tetrazolium salt assays. Cell migration was evaluated using a "scratch assay". Growth factor expression in cells was analyzed via quantitative real-time reverse transcription polymerase chain reaction. RESULTS.: Against the hDFa cell line, the extract had an IC50 of 200 µg/mL and EC50 of 103.5 µg/mL. In the proliferation assay, protein K2 (obtained from the extract) exhibited increased proliferative activity relative to other treatments (1 µg/mL); this agent also exhibited increased activity (50 µg/mL) in the fibroblast migration assay.Furthermore, the relative expression of platelet-derived growth factor increased by 8.6-fold in the presence of K2 protein relative to the control. CONCLUSIONS.: The hydroethanolic extract of Piper aduncum and its component proteins increased the proliferation and migration of hDFa and increased the expression of growth factors involved in the healing process.

Determinación del efecto cicatrizante de Piper aduncum (matico) en fibroblastos humanos / Determination of the healing effect of Piper aduncum (spiked pepper or matico) on human fibroblasts

RESUMEN Objetivos. Evaluar el efecto cicatrizante del extracto hidroetanólico de Piper aduncum, en una línea celular de fibroblastos Dermales Adultos Humanos (hDFa). Materiales y métodos. El extracto se obtuvo mediante extracción sólido-líquido, fue concentrado y liofilizado. Se purificaron las proteínas del extracto mediante cromatografía líquida de alta eficacia de fase reversa (RP-HPLC); las proteínas fueron identificadas por espectrometría de masas en tándem de péptidos trípticos y se analizaron por MALDI-TOF-TOF en un espectrómetro de masa ABSciex4800. Los valores de concentración efectiva media (EC50), concentración inhibitoria media (IC50), y el porcentaje de proliferación celular; fueron determinados por ensayos con sales de tetrazolio (MTT) . La migración celular se evaluó mediante la "técnica de rayado" . Se analizó la expresión de factores de crecimiento mediante el ensayo de reacción en cadena de la polimerasa con transcriptasa reversa a tiempo real (RT- qPCR). Resultados. La línea hDFa evidenció un IC50 de 200 µg/mL con el extracto, el valor de EC50 fue 103,5 µg/mL. En el ensayo de proliferación, la proteína K2; mostró mayor actividad en la proliferación respecto de otros tratamientos (1 µg/mL). En el ensayo de migración de fibroblastos, la proteína K2 mostró mayor actividad (50 µg/mL). La expresión relativa del factor de crecimiento derivado de plaquetas (PDGF) se incrementó 8,6 veces respecto al control, en presencia de la proteína K2. Conclusiones. El extracto hidroetanólico, de Piper aduncum, así como las proteínas que contiene, incrementaron la proliferación y migración de fibroblastos dermales humanos (hDFa); así mismo, aumentaron la expresión de factores de crecimiento que intervienen en el proceso de cicatrización.

ABSTRACT Objectives. To evaluate the healing effect of a Piper aduncum ethanol-water extract on an adult human dermal fibroblast cell line (hDFa). Materials and Methods. After obtaining the extract via solid-liquid extraction, concentration, and lyophilization, extract proteins were purified using reverse phase high-performance liquid chromatography, identified using tandem mass spectrometry of tryptic peptides, and analyzed using MALDI-TOF-TOF on an ABSciex4800 mass spectrometer. Half maximum effective concentration values (EC50), half maximum inhibiting concentration (IC50), and percentages of cell proliferation were determined using tetrazolium salt assays. Cell migration was evaluated using a "scratch assay". Growth factor expression in cells was analyzed via quantitative real-time reverse transcription polymerase chain reaction. Results. Against the hDFa cell line, the extract had an IC50 of 200 μg/mL and EC50 of 103.5 µg/mL. In the proliferation assay, protein K2 (obtained from the extract) exhibited increased proliferative activity relative to other treatments (1 µg/mL); this agent also exhibited increased activity (50 µg/mL) in the fibroblast migration assay.Furthermore, the relative expression of platelet-derived growth factor increased by 8.6-fold in the presence of K2 protein relative to the control. Conclusions. The hydroethanolic extract of Piper aduncum and its component proteins increased the proliferation and migration of hDFa and increased the expression of growth factors involved in the healing process.

PIK3CA mutations in Peruvian patients with HER2-amplified and triple negative non-metastatic breast cancers.

PURPOSE: To determine the frequency of PIK3CA mutations in a Peruvian cohort with HER2-amplified and triple negative breast cancers (TNBC). METHODS: We analyzed two cohorts of 134 primary non-metastatic breast cancer patients from Peru. Cohorts consisted of 51 hormone receptors (+)/HER2-amplified breast tumor patients surgically resected as first treatment included in the ALTTO trial (ALTTO cohort) and 81 TNBC patients with residual disease after neoadjuvant treatment (neoadjuvant cohort). Genomic DNA was extracted from paraffin-embedded tumor samples. Samples from the ALTTO and neoadjuvant cohorts were taken at biopsies and from residual tumors, respectively. PIK3CA mutations were detected by sequencing DNA fragments obtained by PCR amplification of exons and their flanking introns. All of the detected PIK3CA mutations were confirmed in a second independent run of sample testing. RESULTS: PIK3CA mutations were present in 21/134 cases (15.7%). Mutations in exon 9 and 20 were present in 10/134 (7.5%) and 11/134 (8.2%), respectively. No cases had mutations in both exons. Mutations in exon 9 consisted of E545A (seven cases), E545K (two cases) and E545Q (one case); while in exon 20, mutations consisted of H1047R (10 cases) and H1047L (one case). Compared to TNBC patients, HER2-amplified patients were more likely to have PIK3CA mutated (23% vs 9.6%; P=0.034). There were no associations between mutational status of PIK3CA with estrogen receptor status (P=0.731), progesterone receptor status (P=0.921), age (P=0.646), nodal status (P=0.240) or histological grade (P=1.00). No significant associations were found between PIK3CA mutational status and clinicopathological features. CONCLUSIONS: We found a similar frequency of PIK3CA mutations to that reported in other series. Although we did not include HR+/HER2 patients, those with HER2-amplified tumors were more likely to present PIK3CA mutations compared to patients with triple negative tumors.

[Gastrointestinal Epstein Barr virus positive diffuse large B cell lymphoma in elderly: report of five cases and review of literature]. / Linfoma de celulas grandes B difuso gastrointestinal del anciano asociado al virus Epstein Barr: reporte de cinco casos y revision de la literatura.

EBV-positive diffuse large B-cell lymphoma (DLBCL) in elderly is a new entity included provisionally in the most recent WHO Classification of lymphoid neoplasms. It usually affects elderly patients and has a poor survival. The goal of this report was to evaluate clinical, endoscopic characteristics and survival of five patients with this entity and gastrointestinal afectation. From five cases, three cases had gastric infiltration, one ileon and one in cecum.

EBV-positive diffuse large B-cell lymphoma of the elderly: a case series from Peru.

EBV-positive diffuse large B-cell lymphoma (DLBCL) of the elderly is an entity recently included in the WHO classification of lymphoid tumors. We have reviewed our experience and clinical outcomes of this distinct subtype of DLBCL. Between 2002 and 2009, cases of DLBCL were identified from medical records of the Hospital Nacional Edgardo Rebagliati Martins in Lima, Peru, and underwent pathological evaluation including immunohistochemistry for CD20, CD10, bcl-6, MUM1/IRF4, and EBV-encoded RNA in situ hybridization. Clinical data were gathered, tabulated, and reported descriptively. Survival analyses were performed using Kaplan-Meier estimates. Out of 199 cases of DLBCL, 28 cases of EBV-positive DLBCL of the elderly were identified. The median age was 75 years with male predominance (1.5:1). B-symptoms were present in 43%, advanced stage in 50% and International Prognostic Index (IPI) score > 2 in 57% of patients; 68% of patients had a nongerminal center (NGC) phenotype. The complete response rates to R-CHOP and CHOP were 63% and 33%, respectively. The median overall survival (OS) for the group was 5 months. In the univariate analysis, age ≥70 years, lymphocyte count <1.0 × 10(9) /L, and advanced clinical stage were associated with worse OS in patients treated with chemotherapy with and without rituximab. EBV-positive DLBCL of the elderly is a clinically aggressive entity with a short OS and typically presents with advanced stage, high IPI score, and a NGC phenotype. Further studies are needed to investigate if rituximab-containing regimens are associated with better response and OS rates in EBV-positive DLBCL of the elderly.

Linfoma de Celulas Grandes B Difuso Gastrointestinal del Anciano Asociado al Virus Epstein Barr: Reporte de Cinco Casos y Revision de la Literatura / Gastrointestinal Epstein Barr Virus positive diffuse large B cell lymphoma in elderly: report of five cases and review of literatura

El Linfoma de Células Grandes B difuso del anciano asociado al Epstein Barr es una nueva entidad incluida provisionalmente en la más reciente clasificación de neoplasias linfoides de la WHO. Usualmente afecta ancianos y tiene pobre sobrevida. El objetivo de este reporte fue evaluar las característica clínicas, endoscópicas y sobrevida de cinco pacientes portadores de esta entidad y compromiso gastrointestinal. Tres casos tuvieron infiltración gástrica y dos casos tuvieron compromiso del ileón y el ciego.

EBV-positive diffuse large B-cell lymphoma (DLBCL) in elderly is a new entity included provisionally in the most recent WHO Classification of lymphoid neoplasms. It usually affects elderly patients and has a poor survival. The goal of this report was to evaluate clinical, endoscopic characteristics and survival of five patients with this entity and gastrointestinal afectation. From five cases, three cases had gastric infiltration, one ileon and one in cecum.

Deficiency of antigen-presenting cell invariant chain reduces atherosclerosis in mice.

BACKGROUND: Adaptive immunity and innate immunity play important roles in atherogenesis. Invariant chain (CD74) mediates antigen-presenting cell antigen presentation and T-cell activation. This study tested the hypothesis that CD74-deficient mice have reduced numbers of active T cells and resist atherogenesis. METHODS AND RESULTS: In low-density lipoprotein receptor-deficient (Ldlr(-/-)) mice, CD74 deficiency (Ldlr(-/-)Cd74(-/-)) significantly reduced atherosclerosis and CD25(+)-activated T cells in the atheromata. Although Ldlr(-/-)Cd74(-/-) mice had decreased levels of plasma immunoglobulin (Ig) G1, IgG2b, and IgG2c against malondialdehyde-modified LDL (MDA-LDL), presumably as a result of impaired antigen-presenting cell function, Ldlr(-/-)Cd74(-/-) mice showed higher levels of anti-MDA-LDL IgM and IgG3. After immunization with MDA-LDL, Ldlr(-/-)Cd74(-/-) mice had lower levels of all anti-MDA-LDL Ig isotypes compared with Ldlr(-/-) mice. As anticipated, only Ldlr(-/-) splenocytes responded to in vitro stimulation with MDA-LDL, producing Th1/Th2 cytokines. Heat shock protein-65 immunization enhanced atherogenesis in Ldlr(-/-) mice, but Ldlr(-/-) Cd74(-/-) mice remained protected. Compared with Ldlr(-/-) mice, Ldlr(-/-)Cd74(-/-) mice had higher anti-MDA-LDL autoantibody titers, fewer lesion CD25(+)-activated T cells, impaired release of Th1/Th2 cytokines from antigen-presenting cells after heat shock protein-65 stimulation, and reduced levels of all plasma anti-heat shock protein-65 Ig isotypes. Cytofluorimetry of splenocytes and peritoneal cavity cells of MDA-LDL- or heat shock protein-65-immunized mice showed increased percentages of autoantibody-producing marginal zone B and B-1 cells in Ldlr(-/-)Cd74(-/-) mice compared with Ldlr(-/-) mice. CONCLUSIONS: Invariant chain deficiency in Ldlr(-/-) mice reduced atherosclerosis. This finding was associated with an impaired adaptive immune response to disease-specific antigens. Concomitantly, an unexpected increase in the number of innate-like peripheral B-1 cell populations occurred, resulting in increased IgM/IgG3 titers to the oxidation-specific epitopes.

Cystatin C deficiency promotes inflammation in angiotensin II-induced abdominal aortic aneurisms in atherosclerotic mice.

An imbalance between cysteinyl cathepsins and their principal endogenous inhibitor cystatin C (CystC) may favor proteolysis in the pathogenesis of human abdominal aortic aneurysms (AAA), yet a direct role of CystC in AAA remains unproven. This study used CystC and apolipoprotein E (ApoE) compound mutant (CystC(-/-)ApoE(-/-)) mice to examine directly the role of cysteine protease/protease inhibitor imbalance in AAA formation in angiotensin II-induced AAA. CystC-deficiency increased lumenal diameter and lesion size compared with control mice. CystC(-/-) ApoE(-/-) lesions also demonstrated enhanced inflammatory cell accumulation, more severe elastin fragmentation, and fewer smooth muscle cells in the tunica media. Macrophage content, measured as percent positive area (23.2 +/- 1.4% versus 11.2 +/- 1.4%; P = 0.0003) and number of the CD4(+) T cells (ninefold; P = 0.048), increased significantly in CystC(-/-)ApoE(-/-) lesions. CystC deficiency increased cathepsin activity (5.5 fold; P = 0.001) in AAA, yielding greater elastin degradation and proangiogenic laminin-5 gamma2 peptide production, which may account for increased microvascularization in CystC(-/-)ApoE(-/-) compared with ApoE(-/-) lesions. Increased leukocyte adhesion molecule VCAM-1 expression and leukocyte proliferation might also promote inflammation in CystC-deficient AAA. These data indicate that CystC contributes to experimental AAA pathogenesis and that enhanced cysteine protease activity, due to the lack of CystC, favors inflammation in AAA lesions induced in atherosclerotic mice by promoting microvascularization and smooth muscle cell apoptosis as well as leukocytes adhesion and proliferation.

CC chemokine receptor-1 activates intimal smooth muscle-like cells in graft arterial disease.

BACKGROUND: Graft arterial disease (GAD) limits long-term solid-organ allograft survival. The thickened intima in GAD contains smooth muscle-like cells (SMLCs), leukocytes, and extracellular matrix. The intimal SMLCs in mouse GAD lesions differ from medial smooth muscle cells in their function and phenotype. Although intimal SMLCs may originate by migration and modulation of donor medial cells or by recruitment of host-derived precursors, the mechanisms that underlie their localization within grafts and the factors that drive these processes remain unclear. METHODS AND RESULTS: This study of aortic transplantation in mice demonstrated an important function for chemokines beyond their traditional role in leukocyte recruitment and activation. Intimal SMLCs, but not medial smooth muscle cells, express functional CC chemokine receptor-1 (CCR1) and respond to RANTES by increased migration and proliferation. Although RANTES infusion in vivo promoted inflammatory cell accumulation in the adventitia of aortic allografts of wild-type and CCR1-deficient recipients, it increased GAD intimal thickening with SMLC proliferation in only the wild-type hosts. Aortic allografts transplanted into CCR1-deficient mice after wild-type bone marrow transplantation did not develop intimal lesions, which indicates that CCR1-bearing inflammatory cells do not contribute to intimal lesion formation. Moreover, RANTES induced SMLC proliferation in vitro but did not promote medial smooth muscle cell growth. Blockade of CCR5 attenuated RANTES-induced T-cell and monocyte/macrophage proliferation but did not affect RANTES-induced SMLC proliferation, consistent with a larger role of CCR1-binding chemokines in SMLC migration and proliferation and GAD development. CONCLUSIONS: These studies provide a novel mechanistic insight into the formation of vascular intimal hyperplasia and suggest a novel therapeutic strategy for preventing allograft arteriopathy.