Expression of Bcl-2 by human bone marrow mast cells and its overexpression in mast cell leukemia

Bcl-2 protein plays a major role in the prevention of programmed cell death of differentiating cells. In the present study, the expression of cytoplasmic bcl-2 by human Bone Marrow Mast Cells (BMMC) from both normal and pathological bone marrow samples was examined. A total of 35 subjects corresponding to 9 healthy volunteers, 8 cases of adult indolent systemic mast cell disease (SMCD), 4 cases of pediatric mastocytosis (PM), 11 cases of hematological malignancies (HM), 2 cases of reactive bone marrow, and 1 case of mast cell leukemia (MCL) were analyzed. The expression of bcl-2 was studied using quantitative three-color flow cytometry. We also studied the molecular configuration of the bcl-2 gene and other relatives by Southern blot and polymerase chain reaction (PCR) in the MCL case. Bcl-2 expression was detected in BMMC from all samples analyzed. No significant differences on the expression of bcl-2 were detected between BMMC from healthy subjects and patients with SMCD, PM, HM, and reactive bone marrow. By contrast, bcl-2 protein was overexpressed in BMMC from MCL patient without gene rearrangement. Our results show that bcl-2 protein was constitutively expressed by BMMC. BMMC from MCL display overexpression of bcl-2, which could not be related to molecular rearrangements involving the bcl-2 gene. The expression of this protein by mature MC may play a role in the prevention of MC apoptosis and thus help to explain the long survival of these cells. The overexpression of bcl-2 by BMMC in MCL may help to explain their resistance to chemotherapy-induced apoptosis.     

Somatic Alterations Contributing to Metastasis of a Castration‐Resistant Prostate Cancer

Metastatic castration‐resistant prostate cancer (mCRPC) is a lethal disease, and molecular markers that differentiate indolent from aggressive subtypes are needed. We sequenced the exomes of five metastatic tumors and healthy kidney tissue from an index case with mCRPC to identify lesions associated with disease progression and metastasis. An Ashkenazi Jewish (AJ) germline founder mutation, del185AG in BRCA1, was observed and AJ ancestry was confirmed. Sixty‐two somatic variants altered proteins in tumors, including cancer‐associated genes, TMPRSS2‐ERG, PBRM1, and TET2. The majority (n = 53) of somatic variants were present in all metastases and only a subset (n = 31) was observed in the primary tumor. Integrating tumor next‐generation sequencing and DNA copy number showed somatic loss of BRCA1 and TMPRSS2‐ERG. We sequenced 19 genes with deleterious mutations in the index case in additional mCRPC samples and detected a frameshift, two somatic missense alterations, tumor loss of heterozygosity, and combinations of germline missense SNPs in TET2. In summary, genetic analysis of metastases from an index case permitted us to infer a chronology for the clonal spread of disease based on sequential accrual of somatic lesions. The role of TET2 in mCRPC deserves additional analysis and may define a subset of metastatic disease.     

Phage phi29 protein p56 prevents viral DNA replication impairment caused by uracil excision activity of uracil-DNA glycosylase

Protein p56 encoded by the Bacillus subtilis phage 29 inhibits host uracil-DNA glycosylase (UDG) activity. In previous studies, we suggested that this inhibition is likely a defense mechanism developed by phage 29 to prevent the action of UDG if uracilation occurs in DNA either from deamination of cytosine or the incorporation of dUMP during viral DNA replication. In this work, we analyzed the ability of 29 DNA polymerase to insert dUMP into DNA. Primer extension analysis showed that viral DNA polymerase incorporates dU opposite dA with a catalytic efficiency only 2-fold lower than that for dT. Using the 29 DNA amplification system, we found that 29 DNA polymerase is also able to carry out the extension of the dA:dUMP pair and replicate past uracil. Additionally, UDG and apurinic-apyrimidinic endonuclease treatment of viral DNA isolated from 29-infected cells revealed that uracil residues arise in 29 DNA during replication, probably as a result of misincorporation of dUMP by the 29 DNA polymerase. On the other hand, the action of UDG on uracil-containing 29 DNA impaired in vitro viral DNA replication, which was prevented by the presence of protein p56. Furthermore, transfection activity of uracil-containing 29 DNA was significantly higher in cells that constitutively synthesized p56 than in cells lacking this protein. Thus, our data support a model in which protein p56 ensures an efficient viral DNA replication, preventing the deleterious effect caused by UDG when it eliminates uracil residues present in the 29 genome.     

Comparative effect of human and Trypanosoma cruzi calreticulin in wound healing

In orthopaedics, the use of factors that enhance granulation tissue formation and prevent or delay new bone regeneration is sometimes desirable. Calreticulin (CRT), a unique endoplasmic reticulum luminal Ca²⁺‐binding chaperone widely distributed in eukaryotic cells, is involved in many cellular functions. Among them, CRT has an important influence in cutaneous wound healing and diverse processes associated with cutaneous repair, inhibition of angiogenesis, promotion of cell adhesion and antitumour effect. One of the molecules involved in several aspects of the host–parasite interplay is Trypanosoma cruzi calreticulin (TcCRT), which is highly homologous to human calreticulin (HuCRT). Here, recombinant (r)HuCRT and rTcCRT are compared on their abilities to affect fibroblast behaviour in a scratch plate assay, and wound healing in in vivo skin rat models. In molar terms, rTcCRT is three orders of magnitude more efficient than rHuCRT in increasing proliferation and migration of human fibroblasts in vitro. A similar effect was observed in vivo on rat skin wounds and inhibition of bone gap bridging in rabbit unicortical bone osteotomies. Copyright © 2012 John Wiley & Sons, Ltd.     

Comprehensive Treatment of Periorbital Region with Hyaluronic Acid

The periorbital subunit is one of the first facial regions to show signs of aging, primarily due to volume depletion of the soft tissue and bony resorption. Surgical and office-based nonsurgical procedures form an important basis for periorbital rejuvenation. It is important to make a detailed clinical evaluation of the patient to indicate the most appropriate procedure to be performed. With the objective of showing a nonsurgical procedure for the rejuvenation of the periorbital area, the authors describe a technique of applying fillers in the upper and lower periorbital regions, paying attention to the anatomy of this facial region and the type of product to be used besides the expected results of the procedure and its possible adverse effects and complications. The nonsurgical rejuvenation of the periorbicular region with hyaluronic acid is a new and innovative technique. In the opinion of the authors, it is a great aesthetic impact area and consequently brings high satisfaction to patients.Maintaining a youthful and pleasant appearance of the face in today’s culture impacts quality of life in many patients. The facial contour remodeling is being revolutionized by new nonsurgical techniques.Facial aging is a complex and dynamic process. All people age differently as a result of imbalance, disharmony, and disproportion of the aging process between the overlying soft tissue and the underlying bony frameworks.1 The upper periorbital subunit is one of the first facial regions to show signs of aging, and even minor changes in its structure and volume can distort the perceived emotions and health of patients.2 An aesthetic and youthful upper periorbital subunit is characterized by a well-defined brow of appropriate height and shape, fullness of the upper periorbit, a crisp and well-defined upper eyelid crease, minimal skin excess, and good skin quality.3,4In the aging process of this facial area, one group of patients displays signs of aging due predominantly to soft tissue ptosis of the upper eyelid, requiring surgical excision. Another group presents with volume depletion of the soft tissue and bony resorption of the orbit. The loss of septal support, leading to brow prolapse and an exacerbation of upper eyelid fullness and congestion, can also contribute to the aging process. This appearance is characterized by deflation of the upper eyelid as well as hollowing and visibility of the supraorbital bony rim, leading to a sunken, hollow, and skeletonized orbit, which can make the patient appear sickly, anorexic, and old. In recent years, modern facial rejuvenation surgery has evolved toward volume restoration in addition to tissue suspension.2Comprehensive analysis of both soft tissue and bony structural changes are essential for the periorbital rejuvenation. Surgical procedures and office-based nonsurgical procedures form an important basis for periorbital rejuvenation, including cosmeceuticals, chemical peels, laser and light treatments, neurotoxins, and fillers. Improved understanding of the pathophysiology of aging and technical advancements in nonsurgical techniques has enabled us to achieve better and more comprehensive improvement for patients.5     

Delayed-type Necrosis after Soft-tissue Augmentation with Hyaluronic Acid

The growing use of dermal fillers, specifically the use of hyaluronic acid, can be explained by their effectiveness and versatility as well as their favorable safety profiles. Nevertheless, early and late complications with varying levels of severity may occur. The incidence of complications is low and the majority of adverse events are mild (edema, erythema, and local ecchymosis) and of limited duration. However, more severe events, such as ischemia and necrosis, may occur. The symptoms of ischemia can occur immediately after the injection or several hours after the procedure. Here, the authors report three cases of necrosis after hyaluronic acid injection with the first symptoms presenting only several hours after the procedure. The patients were treated immediately after the diagnosis. The aim of this review is to communicate the possibility of the delayed-type presentation of necrosis, present the signs and symptoms that lead to early diagnosis, and review the treatment possibilities of this severe complication.Dermal fillers have been injected with increasing frequency over the past three decades for soft-tissue augmentation by volume expansion in the management of the aging face. In 2012, there were about two million procedures using dermal fillers, according to the American Society of Plastic Surgeons, five percent more than in 2011 and 205 percent more than in 2000, second only to botulinum toxin type A. These minimally invasive and nonsurgical cosmetic procedures were the two most commonly performed in this range of time studied.1,2The growing use of dermal fillers, specifically the use of hyaluronic acid (HA), can be explained by their effectiveness and versatility as well as their favorable safety profiles. Nevertheless, early and late complications with varying levels of severity may occur. The incidence of complications is low and the majority of adverse events are mild (edema, erythema, and local ecchymosis) and of limited duration. However, more severe events, such as ischemia and necrosis, may occur.Injection necrosis is a rare, but important, complication associated with dermal fillers. Necrosis can be attributed to one of two factors—an interruption of vascular supply due to compression or frank obstruction of vessels by direct injection of the material into a vessel itself. The glabella is the injection site commonly believed to be at greater risk for necrosis, but it can also occur at the nasolabial fold.3 Risk factors for intravascular injection include site of application (deep injection of filler products at or near the site of named vessels), volume applied (larger amounts of product can cause a proportionally greater degree of arterial obstruction), and previous scarring (deep tissue scars may stabilize and fix arteries in place, making them easier to penetrate with small sharp needles).4The initial presentation of vascular events may include pain and discomfort disproportionate to what is typically experienced following filler treatments and clinical findings, including blanching, livedo pattern, or violaceous discoloration.4 Although many cases report this immediate post-injection presentation as the typical background seen in a necrosis event, there are few reports with the first symptom presenting only hours after augmentation. See Figures 1 through ​through3,3, where the authors present three cases of vascular compromise after soft-tissue augmentation with delayed-type presentation. Open in a separate windowOpen in a separate windowFigures 2Aand 2B.Case 2: Necrosis and secondary infection 48 hours after the HA injection (a). Discrete scars in the affected area after treatment (b). Open in a separate windowOpen in a separate windowFigures 1Aand 1B.Case 1: Edema, erythema, and progressive violaceous reticulated patch, livedoid area were observed on the left cheek 36 hours after the injection (a). Complete healing five days after hyaluronidase application and nine days after the HA injection (b). Open in a separate windowOpen in a separate windowFigures 3Aand 3B.Case 3: Necrosis and secondary infection 48 hours after the HA injection (a). Erythema, hipercromia, and discreet scars in the affected area after treatment (b).