José Vassallo

Loss-of-Function Mutation in Toll-Like Receptor 4 Prevents Diet-Induced Obesity and Insulin Resistance

Obesity is associated with insulin resistance and a state of abnormal inflammatory response. The Toll-like receptor (TLR)4 has an important role in inflammation and immunity, and its expression has been reported in most tissues of the body, including the insulin-sensitive ones. Because it is activated by lipopolysaccharide and saturated fatty acids, which are inducers of insulin resistance, TLR4 may be a candidate for participation in the cross-talk between inflammatory and metabolic signals. Here, we show that C3H/HeJ mice, which have a loss-of-function mutation in TLR4, are protected against the development of diet-induced obesity. In addition, these mice demonstrate decreased adiposity, increased oxygen consumption, a decreased respiratory exchange ratio, improved insulin sensitivity, and enhanced insulin-signaling capacity in adipose tissue, muscle, and liver compared with control mice during high-fat feeding. Moreover, in these tissues, control mice fed a high-fat diet show an increase in IκB kinase complex and c-Jun NH2-terminal kinase activity, which is prevented in C3H/HeJ mice. In isolated muscles from C3H/HeJ mice, protection from saturated fatty acid–induced insulin resistance is observed. Thus, TLR4 appears to be an important mediator of obesity and insulin resistance and a potential target for the therapy of these highly prevalent medical conditions.

An inherited mutation leading to production of only the short isoform of GATA-1 is associated with impaired erythropoiesis

Acquired somatic mutations in exon 2 of the hematopoietic transcription factor GATA-1 have been found in individuals with Down syndrome with both transient myeloproliferative disorder and acute megakaryoblastic leukemia. These mutations prevent the synthesis of the full-length protein but allow the synthesis of its short isoform, GATA-1s. Experiments in mice suggest that GATA-1s supports normal adult megakaryopoiesis, platelet formation and erythropoiesis. Here we report a mutation, 332G → C, in exon 2 of GATA1, leading to the synthesis of only the short isoform in seven affected males from two generations of a family. Hematological profiles of affected males demonstrate macrocytic anemia, normal platelet counts and neutropenia in most cases. Altogether, data suggest that GATA-1s alone, produced in low or normal levels, is not sufficient to support normal erythropoiesis. Moreover, this is the first study to indicate that a germline splicing mutation does not lead to leukemia in the absence of other cooperating events, such as Down syndrome.

Digital slides: present status of a tool for consultation, teaching, and quality control in pathology.

In the last few years, telepathology has benefited from the progress in the technology of image digitalization and transmission through the world web. The applications of telepathology and virtual imaging are more current in research and morphology teaching. In surgical pathology daily practice, this technology still has limits and is more often used for case consultation. In the present review, we intend to discuss its applications and challenges for pathologists and scientists. Much of the limitations of virtual imaging for the surgical pathologist reside in the capacity of storage of images, which so far has hindered the more widespread use of this technology. Overcoming this major drawback may revolutionize the surgical pathologists activity and slide storing.

ALK-positive anaplastic large cell lymphoma mimicking nodular sclerosis Hodgkin's lymphoma: Report of 10 cases

Anaplastic large cell lymphoma (ALCL) and Hodgkin lymphoma (HL) are recognized as biologically distinct entities. However, occasionally, these two entities may share some morphologic features responsible for diagnostic difficulties. In the last 10 years, we have collected 380 cases of ALK-positive ALCL of which 10 cases were originally diagnosed as nodular sclerosis classic HL (NSHL) on conventional histopathological examination. After immunostaining, these cases proved to be ALK-positive ALCL mimicking HL (so-called Hodgkin-like ALCL). These cases account for 2.6% of our cases of ALK-positive ALCL (10 of 380 cases). Median age was 11 years (3-92 years) with a female predominance (male/female ratio, 3:7). Characteristically, these lesions showed thick nodular fibrosing bands highly suggestive of NSHL. Neoplastic cells were scarce in 6 cases, whereas in the 4 remaining cases, sheets of tumor cells were also present. A perivascular and a sinusoidal growth pattern was observed in various degrees in all cases. Few binucleated Reed-Sternberg-like cells were present in every case in a background of small lymphocytes. Inflammatory cells (ie, granulocytes, eosinophils, and histiocytes) were rare. Neoplastic cells were positive for CD30 (10 of 10 cases), ALK protein (10 of 10 cases), epithelial membrane antigen (EMA) (9 of 9 cases), CD43 (6 of 9 cases), and perforin (8 of 8 cases), but negative for CD15 (10 of 10 cases), CD20 (10 of 10 cases), Pax5/BSAP (6 of 6 cases), and EBV (8 of 8 cases). In addition, in 7 cases, neoplastic cells were of T-phenotype, whereas the 3 remaining cases were considered to be of null/undetermined phenotype. Although rare, Hodgkin-like ALCL may mimic NSHL, and it is advisable to include EMA in the first line panel and to ask for ALK staining in EMA-positive, CD15-negative lesions with morphologic features suggestive of NSHL.

EGFR Tyrosine Kinase Inhibitor (PD153035) Improves Glucose Tolerance and Insulin Action in High-Fat Diet–Fed Mice

OBJECTIVE In obesity, an increased macrophage infiltration in adipose tissue occurs, contributing to low-grade inflammation and insulin resistance. Epidermal growth factor receptor (EGFR) mediates both chemotaxis and proliferation in monocytes and macrophages. However, the role of EGFR inhibitors in this subclinical inflammation has not yet been investigated. We investigated, herein, in vivo efficacy and associated molecular mechanisms by which PD153035, an EGFR tyrosine kinase inhibitor, improved diabetes control and insulin action. RESEARCH DESIGN AND METHODS The effect of PD153035 was investigated on insulin sensitivity, insulin signaling, and c-Jun NH2-terminal kinase (JNK) and nuclear factor (NF)-κB activity in tissues of high-fat diet (HFD)-fed mice and also on infiltration and the activation state of adipose tissue macrophages (ATMs) in these mice. RESULTS PD153035 treatment for 1 day decreased the protein expression of inducible nitric oxide synthase, tumor necrosis factor (TNF)-α, and interleukin (IL)-6 in the stroma vascular fraction, suggesting that this drug reduces the M1 proinflammatory state in ATMs, as an initial effect, in turn reducing the circulating levels of TNF-α and IL-6, and initiating an improvement in insulin signaling and sensitivity. After 14 days of drug administration, there was a marked improvement in glucose tolerance; a reduction in insulin resistance; a reduction in macrophage infiltration in adipose tissue and in TNF-α, IL-6, and free fatty acids; accompanied by an improvement in insulin signaling in liver, muscle, and adipose tissue; and also a decrease in insulin receptor substrate-1 Ser307 phosphorylation in JNK and inhibitor of NF-κB kinase (IKKβ) activation in these tissues. CONCLUSIONS Treatment with PD153035 improves glucose tolerance, insulin sensitivity, and signaling and reduces subclinical inflammation in HFD-fed mice.

Prognostic significance of PD-L1 and PD-L2 in breast cancer

PD-L1 and PD-L2 constitute an important antitumor immune response. In breast cancer, their prognostic value is still to be defined. In this study, we investigate the correlation between PD-L1 and PD-L2 protein expressions with clinical and pathologic features and disease-free survival and overall survival. To assess PD-L1 and PD-L2 expressions, we conducted immunohistochemistry studies using a breast cancer tissue microarray encompassing a total of 192 breast cancer cases, stages I, II, and III, with detailed clinical and outcome data. PD-L1 expression was present in 56.6% (107/189), and PD-L2 expression was identified in 50.8% (97/191) of breast cancer cases. Younger age at diagnosis, lymph node positivity, negative estrogen receptor, and recurrence at distant sites were all associated with both PD-L1 and PD-L2 expressions. The presence of larger tumors was associated only with PD-L1 expression. In our study, PD-L1 expression was significantly associated with better overall survival (P = .04) in breast cancer patients. Despite its association with poor clinical and pathologic features, PD-L1 expression emerges as a positive prognostic biomarker in breast cancer. This survival result might be due to the presence of a strong antitumor immune response leading to PD-L1 expression.

Infiltration of a mixture of immune cells may be related to good prognosis in patients with differentiated thyroid carcinoma

Immune responses against differentiated thyroid carcinomas (DTC) have long been recognized. We aimed to investigate the role of immune cell infiltration in the progression of DTC.

Clinical and Pathological Implications of Concurrent Autoimmune Thyroid Disorders and Papillary Thyroid Cancer

Cooccurrences of chronic lymphocytic thyroiditis (CLT) and thyroid cancer (DTC) have been repeatedly reported. Both CLT and DTC, mainly papillary thyroid carcinoma (PTC), share some epidemiological and molecular features. In fact, thyroid lymphocytic inflammatory reaction has been observed in association with PTC at variable frequency, although the precise relationship between the two diseases is still debated. It also remains a matter of debate whether the association with a CLT or even an autoimmune disorder could influence the prognosis of PTC. A better understanding about clinical implications of autoimmunity in concurrent thyroid cancer could raise new insights of thyroid cancer immunotherapy. In addition, elucidating the molecular mechanisms involved in autoimmune disease and concurrent cancer allowed us to identify new therapeutic strategies against thyroid cancer. The objective of this article was to review recent literature on the association of these disorders and its potential significance.

Detection of Epstein-Barr virus in tonsillar tissue of children and the relationship with recurrent tonsillitis.

Recurrent tonsillitis has been the subject of much investigation. Events considered to predispose to or cause recurrent tonsillitis (RT) include the misuse of antibiotic therapy in acute bouts, alterations in the microflora, structural changes in crypt epithelium and certain viral infections. Epstein-Barr Virus (EBV) infection usually occurs in early childhood and can persist in palatine tonsil lymphocytes to induce tonsillitis at a later date. We have examined the presence of EBV in palatine tonsils in order to assess the relationship between this virus and recurrent acute tonsillitis. Tonsils were obtained from 85 patients, 2--14 years old (mean 5.6 years old) who underwent tonsils and adenoid (T&A) removal because of recurrent tonsillitis (RT) or T&A hypertrophy (TH). Tissues specimens were processed for non-isotopic in situ hybridization (ISH) using EBER 1/2 oligonucleotides (EBER RNA). The indications for surgery were RT in 42 patients and TH in 43 patients. In 25 out of 85 cases (29.4%) a positive EBER RNA reaction (15 RT and 33 TH) was found. The chi(2)-test showed no statistically significant difference in frequency of positive results between RT and TH group. We conclude that tonsils of children can be colonized by EBV and that the virus may be implicated in RT and TH.

Model for human skin reconstructed in vitro composed of associated dermis and epidermis

CONTEXT AND OBJECTIVE The technique of obtaining human skin with dermis and epidermis reconstructed from cells isolated from patients can enable autologous skin grafting on patients with few donor sites. It also enables in vitro trials on chemicals and drugs. The objective of this work was to demonstrate a method for obtaining human skin composed of associated dermis and epidermis, reconstructed in vitro. DESIGN AND SETTING Experimental laboratory study, in the Skin Cell Culture Laboratory of Faculdade de Ciências Médicas, Universidade Estadual de Campinas. METHODS Cells from human fibroblast cultures are injected into bovine collagen type I matrix and kept immersed in specific culturing medium for fibroblasts. This enables human dermis reconstruction in vitro. On this, by culturing human keratinocytes and melanocytes, differentiated epidermis is formed, leading to the creation of human skin composed of associated dermis and epidermis, reconstructed in vitro. RESULTS We showed that human skin composed of associated dermis and epidermis can be successfully reconstructed in vitro. It is histologically formed in the same way as human skin in vivo. Collagen tissue can be identified in the dermis, with cells and extracellular matrix organized in parallel to multilayer epidermis. CONCLUSIONS It is possible to obtain completely differentiated human skin composed of associated dermis and epidermis, reconstructed in vitro, from injection of human fibroblasts into bovine collagen type I matrix and culturing of human keratinocytes and melanocytes on this matrix.