Nicotinic acetylcholine receptor agonists may be a novel therapy for endometriosis

Endometriosis (EMs) is defined as the presence of tissue which somewhat resembles endometrial glands and stroma outside the uterus, and elicit an inflammatory response. This response is accompanied by angiogenesis, adhesions, fibrosis, scarring, neuronal infiltration, and anatomical distortion, resulting in pain and infertility. Owing to the side effects of the present medical treatment and the 10% incidence of recurrence after surgery, EMs is difficult to cure completely so far, that have given impetus to consider novel therapy. Since 1980s, cigarette smoking was found inversely related to the risk of having EMs and it is generally considered that nicotine may play a beneficial role in the pathological process of EMs. Recently, the anti-inflammatory and anti-nociceptive functions of nicotinic acetylcholine receptors (nAChRs) as well as the related mechanisms have become a research hotspot. Based on the above-mentioned, it suggests that the nicotinic acetylcholine receptor agonists may be applied for the treatment of EMs.     

Vitamin D receptor agonists as anti-inflammatory agents

1α,25-dihydroxyvitamin D3 (1,25-[OH]2D3), the biologically active form of vitamin D, is a secosteroid hormone that is essential for bone and mineral homeostasis. In addition, this hormone regulates the growth and differentiation of many cell types and has pronounced immunoregulatory and anti-inflammatory properties. Current therapeutic indications include osteoporosis, secondary hyperparathyroidism and psoriasis, but the anti-proliferative, prodifferentiative, antibacterial, immunomodulatory and anti-inflammatory properties of vitamin D receptor agonists could be exploited in a variety of additional clinical conditions. In particular, the pleiotropic anti-inflammatory effects induced by vitamin D receptor agonists could turn out to be beneficial in different pathologies mediated by chronic inflammatory responses.     

The amyloid paradox: amyloid-beta-metal complexes can be neurotoxic and neuroprotective

Senile plaques in the brains of people with Alzheimer disease (AD) are primarily composed of the amyloid-beta (Abeta) peptide and contain substantially elevated levels of iron, copper and zinc. These metals bind to Abeta and have been reported to increase the toxicity of Abeta to cultured neurones. Other reports have demonstrated that Abeta can reduce the neurotoxicity of metal ions, suggesting that the interaction can, under some circumstances, be protective. To investigate these apparently conflicting results, human Abeta1-42 was co-injected with iron, copper or zinc (at the concentrations found in plaques) into rat cerebral cortex, and the resulting numbers of dying neurones were compared. It was found that Abeta complexed with either iron or zinc was more toxic than Abeta alone. In contrast, Abeta-copper complexes were not neurotoxic. Surprisingly, we observed that when iron or copper were combined with Abeta, the neurotoxicity of these metals was substantially reduced, suggesting that Abeta may help to limit the toxicity of redox-active metal ions, thereby assisting the antioxidant defence of the brain. Thus paradoxical effects occur when Abeta complexes with metal ions, where Abeta-metal complexes are capable of being neurotoxic and neuroprotective.     

Overexpressed cell surface interleukin-4 receptor molecules can be successfully targeted for antitumor cytotoxin therapy

A variety of human solid cancer cell lines and primary cell cultures has been reported to overexpress high-affinity receptors (R) for interleukin-4 (IL-4), a pleiotropic immunoregulatory cytokine. The significance of IL-4R expression is not known; however, IL-4 is able to upregulate adhesion molecules, inhibit cell proliferation, and mediate signal transduction in tumor cell lines. To target IL-4R, we produced a chimeric protein composed of a circular permuted IL-4 and a mutated form of Pseudomonas exotoxin [termed IL4(38-37)-PE38KDEL or cpIL4-PE]. The recombinant cpIL4-PE was highly cytotoxic to cancer cells, but not toxic to normal B cells, T cells, monocytes, and CD34+, even though these cells express detectable numbers of IL-4R. The cytotoxicity was specific because excess of recombinant IL-4 neutralized the cpIL4-PE effect. To further develop this molecule, in vivo antitumor activity was tested in animal models of human cancer. This agent showed remarkable antitumor activity in AIDS-Kaposi's sarcoma, glioblastoma multiforme, and breast cancer models in immunodeficient animals. cpIL4-PE caused partial or complete regression of established human tumors. Preclinical efficacy and toxicity studies provided a therapeutic window in which this cancer-targeted agent could be used. On the basis of these studies, we initiated a Phase I clinical trial for the treatment of recurrent glioblastoma multiforme. Our preliminary clinical results suggest that cpIL4-PE has antitumor activity against the deadliest form of brain tumors, without detectable toxicity to normal brain tissues. Thus, IL-4 receptors represent novel targets for cancer cytotoxin therapy.     

Repairing neuronal circuits with brain grafts: Where can brain grafts be used as a therapy?

Transplanted brain tissues are unlikely to become completely integrated into a mature host brain. This places a limitation upon the types of neuronal circuitry that brain grafts can be used to repair. Brain disorders that may be repairable by brain grafts are those with a loss of unidirectional inputs to anatomically restricted brain regions.     

Copper stained protein gels can be efficiently used for immunoblotting.

We have found that the previously described fast and sensitive copper staining of proteins resolved by sodium dodecyl sulphate-polyacrylamide gel electrophoresis does not interfere with the subsequent electrotransfer of these proteins to a solid support and their detection by specific antibodies. After the gel is copper stained and photographed it is simply destained and then equilibrated in transfer buffer prior to immunoblotting. We find that this treatment has no significant effect on transfer efficiency or band sharpness and is compatible with all common detection methods for the blotted proteins. It thus permits the separation of proteins to be checked in a simple way before immunoblotting is performed.     

Glucocorticoid-induced granzyme A expression can be used as a marker of glucocorticoid sensitivity for acute lymphoblastic leukemia therapy

The ability of glucocorticoids (GC) to efficiently kill lymphoid cells has led to their inclusion in essentially all chemotherapy procedures used to treat acute lymphoblastic leukemia (ALL). GC sensitivity is an important prognostic factor in ALL treatment, and it is used to classify patients into risk groups. Clinical assessment for GC sensitivity is very time-consuming, however. We have recently found that granzyme A (GZMA) mediates GC-induced apoptosis in ALL-derived cell line 697. In this study we examined the correlation between GC sensitivity and GC-induced GZMA expression by using seven established cell lines derived from ALL patients. The apoptosis assay showed four cell lines were GC-sensitive and three were GC-resistant. GC treatment markedly enhanced GZMA expression in GC-sensitive cell lines only, and not in GC-resistant cell lines. GC-induced GZMA expression also correlated well with the amount of GC-induced apoptosis. GC-induced GZMA expression could thus be a useful early biomarker for "personalized" ALL therapy.     

Stem cells therapy: a review on approaches that can be used for treatment of respiratory failures in sulfur mustard-injured patients

Abstract

Sulfur mustard (SM) is a toxic agent which causes severe abnormalities in an airway system such as necrosis and inflammation, oxidative stress, chronic bronchitis, shortness of breath, and chronic obstructive pulmonary disease. Although possible mechanisms of SM toxicity have been extensively considered, there is still need to find an appropriate clinical treatment to decrease chronic lung injuries caused by SM. Due to extensive progresses and achievement in tissue repairing through stem cells therapy, the importance of cell therapy for the treatment of lung injuries has been increased. However, several factors such as types of stem cells, necessary conditions for growth and proliferation of stem cells, and their homing into the target tissues are considered as the most important problems in this issue. Mesenchymal stem cells (MSCs) are a class of multipotent stem cells with proliferative and self-renewal capacity which are able to differentiate into different cell lines such as lung epithelial cells. They have a potential repairing and immune modulatory properties which make them as a good candidate for the regeneration of bronchioles tract in SM-exposed patients. Unlike chemical drugs, the differentiation and high-level safety properties of MSCs can be considered as a new strategy for the treatment of SM-injured patients with pulmonary complications. This review aims to consider the therapeutic effects of MSCs in the treatment of SM-induced pulmonary injuries in both animals and humans.     

Tissues other than bone marrow that can be used for cytogenetic analyses

“Sucupira” oil and the lactone eremanthine, extracted from Pterodon pubescens and Eremanthus elaeagnus, respectively, are known for their cercaricidal action in experimental animals. Because of their biological effect, they have the potential to be used for the prophylaxis of schistosomiasis caused by Schistosoma mansoni. To test the clastogenicity of these agents, “sucupira” oil, either pure or diluted in corn oil, was tested in vivo on Wistar rat bone marrow cells following dermal application. Metaphase analysis showed that the compound did not induce a significant increase in the frequencies of chromosomal aberrations. When eremanthine was tested on BALB/c mice following gavage at doses of 100, 200, and 300 mg/kg bw, it did not induce structural or numerical chromosomal aberrations. In the in vitro treatment of human lymphocyte cultures, eremanthine also did not cause any increase in chromosomal aberrations or sister chromatid exchanges at the following concentrations in culture medium: 1.25, 2.50, and 5.00 μg/ml. From these results, under our experimental conditions, neither “sucupira” oil nor eremanthine showed clastogenic effects on mammalian cells in vivo or in vitro. © 1995 Wiley-Liss, Inc.     

Behavioral versus insight-oriented marital therapy: labels can be misleading

Snyder, Wills, and Grady-Fletcher (1991) reported a 4-year follow-up comparing insight-oriented (IOMT) with behavioral marital therapy (BMT). The effects of IOMT were more durable than those of BMT. These comments question the adequacy with which the two treatments were represented. The BMT treatment manual adequately represents behavioral technology as it existed in 1980 but fails to include more recent clinical innovations. The IOMT treatment manual includes many clinical skills that are integral to BMT but not included in the BMT manual. Documentation is provided that therapists in the BMT condition were not using these important techniques. These and other questions regarding the adequacy of training and supervision may have compromised the integrity of BMT. Nevertheless, the findings do suggest that traditional BMT technology alone may be neither necessary nor sufficient for long-term change.     

The allergy-associated chemokine receptors CCR3 and CCR5 can be inactivated by the modified chemokine NNY-CCL11

BACKGROUND: CC chemokine ligand 11 (CCL11) is the outstanding member of all described CC chemokine receptor 3 (CCR3) ligands and is shown to be selective for this receptor. However, it also activates CCR5 but only in the micromolar range. The in vivo activity of CCL11 is expected to be temporally restricted, as it is degraded by specific proteases such as the dipeptidyl-peptidase IV (DP4), also termed CD26. Based on the approach to inactivate chemokine receptors in allergic disease models as has been demonstrated for DP4-resistant n-nonanoyl (NNY)-CCL14 and for amino-oxypentane (AOP)-CCL5, it is tempting to study similar compounds derived from CCL11. METHODS: Synthesis of NNY-CCL11 was performed and it was characterized for biological functions in human and mouse eosinophils as well as in cell lines stably transfected either with human CCR3 or CCR5. Resistance to DP4 treatment was also investigated. RESULTS: The functional activities of NNY-CCL11 mediated via CCR3 show an almost identical pattern to CCL11 with respect to intracellular calcium mobilization and CCR3 internalization. N-terminal cleavage of CCL11 by preincubation with DP4 results in a reduced capacity to internalize CCR3, while preincubation of NNY-CCL11 shows no influence. In contrast to CCL11, NNY-CCL11 also activates CCR5+ cell lines and human monocytes in the nanomolar range, being about 100 times more potent than CCL11. CONCLUSIONS: n-Nonanoyl-CCL11 represents a compound with dual activity restricted to CCR3 and CCR5. Because of its receptor-inactivating capacity and stability against DP4 degradation, NNY-CCL11 is a suitable tool for the decoding of the pathophysiological mechanisms of allergic diseases.     

Self-sampled and air-dried cervicovaginal secretions can be used for analyses of mucosal antibodies

Cervicovaginal secretions were collected from 26 women (13 premenopausal and 13 postmenopausal) using a new sampling device (MucoSafe™) with an absorbent which was introduced into the vagina and retrieved by the women themselves, after which it was air-dried and stored for months at room temperature until extraction of immunoglobulins. Cervical secretions were also collected by absorbent cylindrical wicks (Polyfiltronics) which were introduced into the cervical canal during speculum examination and thereafter kept frozen until extraction. The concentrations of specific IgA and IgG antibodies (to group B streptococci) in extracts from both methods were corrected by reference to total immunoglobulin levels. Three pairs of samples, all from postmenopausal women, were excluded from analysis due to undetectable levels of antibodies in the MucoSafe™ specimen. In the remaining 23 pairs, corrected concentrations of IgA and IgG antibodies in samples obtained by MucoSafe™ correlated well with the corresponding concentrations in wick samples, R=0.84 (p<0.0001) and R=0.69 (p=0.0002), respectively. Thus, cervicovaginal secretions for antibody measurements can be obtained by this novel method for self-sampling, obviating the need for speculum examination and storage of frozen samples.     

Kidney biopsies can be used for estimations of glomerular number and volume: a pig study

In this study, we wanted to evaluate the use of kidney biopsies for estimation of N(glom) and V(glom) in both healthy and chronically diseased kidneys. Danish Landrace pigs with mean weight of 29 kg (range: 25–35 kg) were either subjected to unilateral ureteral obstruction (UUO) or non-obstruction (healthy). N(glom) and V(glom) was estimated by design-based methods using biopsies, N(glom)^biopsy and V(glom)^biopsy. From each kidney, six biopsies were withdrawn at six topographically different sites. All estimates were done following stereological principles and reference methods estimated number with the physical fractionator, N(glom)^PF, and volume with test point system, V(glom)^TPS. N(glom)^PF was for UUO kidneys and for healthy kidneys. N(glom)^biopsy was (p > 0.05) for UUO and (p = 0.04) for healthy kidneys. When UUO and healthy kidneys were grouped, N(glom)^PF was , and N(glom)^biopsy was (p > 0.05). V(glom)^TPS was 1,079 ± 126 mm³ for UUO and 1,707 ± 263 mm³ for healthy kidneys. V(glom)^biopsy was 1,048 ± 291 mm³ for UUO (p > 0.05) and 1,373 ± 393 mm³ for healthy kidneys (p > 0.05). When UUO and healthy kidneys were grouped, V(glom)^TPS was 1,180 ± 229 mm³ and V(glom)^biopsy 1,129 ± 334 mm³ (p > 0.05). Biopsy sites were tested for any systematic differences between site- and mean values, and no significant difference was found (p > 0.05). This study showed that biopsies can be used for estimating N(glom) and V(glom) by design-based methods, but more precise determination of biopsy volume is needed.     

Natural and synthetic agonists of the melanocortin receptor type 3 possess anti-inflammatory properties

The effects of the natural and synthetic ligands for the melanocortin receptor type 3 (MC3-R) have been evaluated in a murine model of experimental gout. Systemic treatment of mice with gamma2-melanocyte-stimulating hormone (gamma2-MSH) and the synthetic agonist MTII inhibited accumulation of KC, interleukin-1 beta (IL-1beta), and PMN elicited by urate crystals in the peritoneal cavity. In vitro, macrophage (M?) activation, determined as release of KC and IL-1beta, was inhibited by gamma2-MSH and MTII. The mixed MC3/4-R antagonist SHU9119 prevented the inhibitory actions of gamma2-MSH and MTII in vitro and in vivo, whereas the selective MC4-R antagonist HS024 was without effect. Western blotting also showed the presence of MC3-R protein on murine peritoneal M?. Furthermore, agonism at the MC3-R evoked accumulation of cAMP within the M?, which was inhibited by SHU9119. Thus, naturally occurring melanocortins, as well as the synthetic long-acting compound MTII, activate MC3-R on peritoneal M? to inhibit the experimental inflammatory response.     

Methodological requirements for valid tissue-based biomarker studies that can be used in clinical practice

Paralleling the growth of ever more cost efficient methods to sequence the whole genome in minute fragments of tissue has been the identification of increasingly numerous molecular abnormalities in cancers—mutations, amplifications, insertions and deletions of genes, and patterns of differential gene expression, i.e., overexpression of growth factors and underexpression of tumor suppressor genes. These abnormalities can be translated into assays to be used in clinical decision making. In general terms, the result of such an assay is subject to a large number of variables regarding the characteristics of the available sample, particularities of the used assay, and the interpretation of the results. This review discusses the effects of these variables on assays of tissue-based biomarkers, classified by macromolecule—DNA, RNA (including micro RNA, messenger RNA, long noncoding RNA, protein, and phosphoprotein). Since the majority of clinically applicable biomarkers are immunohistochemically detectable proteins this review focuses on protein biomarkers. However, the principles outlined are mostly applicable to any other analyte. A variety of preanalytical variables impacts on the results obtained, including analyte stability (which is different for different analytes, i.e., DNA, RNA, or protein), period of warm and of cold ischemia, fixation time, tissue processing, sample storage time, and storage conditions. In addition, assay variables play an important role, including reagent specificity (notably but not uniquely an issue concerning antibodies used in immunohistochemistry), technical components of the assay, quantitation, and assay interpretation. Finally, appropriateness of an assay for clinical application is an important issue. Reference is made to publicly available guidelines to improve on biomarker development in general and requirements for clinical use in particular. Strategic goals are formulated in order to improve on the quality of biomarker reporting, including issues of analyte quality, experimental detail, assay efficiency and precision, and assay appropriateness.