Antibody-Guided In Vivo Imaging for Early Detection of Mammary Gland Tumors

BACKGROUND: Earlier detection of transformed cells using target-specific imaging techniques holds great promise. We have developed TAB 004, a monoclonal antibody highly specific to a protein sequence accessible in the tumor form of MUC1 (tMUC1). We present data assessing both the specificity and sensitivity of TAB 004 in vitro and in genetically engineered mice in vivo. METHODS: Polyoma Middle T Antigen mice were crossed to the human MUC1.Tg mice to generate MMT mice. In MMT mice, mammary gland hyperplasia is observed between 6 and 10 weeks of age that progresses to ductal carcinoma in situ by 12 to 14 weeks and adenocarcinoma by 18 to 24 weeks. Approximately 40% of these mice develop metastasis to the lung and other organs with a tumor evolution that closely mimics human breast cancer progression. Tumor progression was monitored in MMT mice (from ages 8 to 22 weeks) by in vivo imaging following retro-orbital injections of the TAB 004 conjugated to indocyanine green (TAB-ICG). At euthanasia, mammary gland tumors and normal epithelial tissues were collected for further analyses. RESULTS: In vivo imaging following TAB-ICG injection permitted significantly earlier detection of tumors compared with physical examination. Furthermore, TAB-ICG administration in MMT mice enabled the detection of lung metastases while sparing recognition of normal epithelia. CONCLUSIONS: The data highlight the specificity and the sensitivity of the TAB 004 antibody in differentiating normal versus tumor form of MUC1 and its utility as a targeted imaging agent for early detection, tumor monitoring response, as well as potential clinical use for targeted drug delivery.     

A comparative study of myosin and its subunits in adult and neonatal-rat hearts and in rat heart cells from young and old cultures.

A possible explanation for the decrease in myosin Ca2+-dependent ATPase activity as rat heart cells age in culture is presented. The subunit structure and enzyme kinetics of myosin from adult and neonatal rat hearts and from rat heart cells of young and old cultures are compared. These studies indicate that the loss in Ca-ATPase activity of myosin from older cultures was an intrinsic property of the myosin itself. Myofibrillar fractions from the indicated four sources showed no qualitative or quantitative differences in electrophoretic patterns. Myosin from older cultures was more sensitive to alkaline denaturation than was myosin from younger cultures, as indicated by its more accelerated loss of K+(EDTA)-dependent ATPase activity after 10 min of incubation at pH 10. Furthermore, myosin from older cultures was more temperature-sensitive, as indicted by a more rapid loss of Ca-ATPase with decrease in assay temperature. It is suggested that there is either a change in conformation of myosin molecules at or near the active site of the enzyme or alternatively there is a change in light chain 1-light chain 2 and/or light-chain-heavy-chain interaction(s) in the myosin molecules under study.     

The measurement of ecstasy in human hair by triple phase directly suspended droplet microextraction prior to HPLC-DAD analysis

New pre-concentration technique, triple phase suspended droplet microextraction (SD-LPME) and liquid chromatography-photodiode array detection was applied to determine ecstasy, MDMA (3,4-methylendioxy-N-methylamphetamine) in hair samples. In this research MDMA in hair was digested and after treatment extracted. The effective parameters were investigated and method was evaluated. Under the optimal conditions, the MDMA was enriched by factor 98.11. Linearity (r = 0.9921), was obtained in the range of 10–15,000 ng mL^?1 and detection limit was 0.1 ng mL^?1.     

Effectiveness of a tris-based extender (SHOTOR diluent) for the preservation of Bactrian camel (Camelus bactrianus) semen

The development of a suitable semen extender is required to extend artificial breeding programs and to preserve the genetic potential of Bactrian camel. Experiments were conducted to provide the optimal osmolality and pH of tris-based extender and to compare that with available extenders for short-term preservation of Bactrian camel semen at 4 degrees C during 24 h. In experiments I and II, the effects of varying osmolalities (270, 300, 330, 360, and 390 mOsm/kg) and pHs (5.5, 6, 6.9, 7.5, 7.9, and 8.9) of tris-based extender on sperm viability were investigated. In experiment III, the efficiency of tris-based extender (SHOTOR diluent) in preserving Bactrian camel semen was compared with lactose (10%), sucrose (10%) and Green buffer. Viability parameters including progressive forward motility (PFM), plasma membrane integrity and the percentage of live spermatozoa were assessed. The data were analyzed using general linear model procedure. In the majority of assessments using tris-based extender, the viability of spermatozoa was superior at the osmolality of 330 mOsm/kg and pH of 6.9. PFM was significantly greater at the time of semen dilution in tris-based (65.5%) and Green buffer (60.5%) compared to that of lactose (31%) and sucrose (28%) extenders (P<0.05), and remained elevated throughout the experiment. There was no significant difference in other viability parameters among 4 extenders (P>0.05). In conclusion, the utilization of a tris-based extender, having the osmolality of 330 mOsm/kg and pH of 6.9, favors the short-term preservation of the Bactrian camel spermatozoa under chilled condition.     

Differential regulation of glutamate receptors in Alzheimer's disease

Selective neurodegeneration is a prominent feature in Alzheimer's disease; however, the mechanism of neuronal death is still unclear. Nonetheless, the topographical distribution of different types of receptors is thought to contribute to the regional selective nature of neuronal degeneration. Specifically, since glutamatergic transmission is severely altered by the early degeneration of cortico-cortical connections and hippocampal projections in Alzheimer's disease, we suspect that glutamate receptors may play a new role in the pathophysiology of disease. Here we review the salient aspects of glutamate receptor expression in Alzheimer's disease and how their differential regulation can contribute to the selective neurodegeneration seen in the disease. Additionally, we assess the potential therapeutic value of glutamate receptors as a target for drug intervention in Alzheimer's disease.     

Erythrocyte membrane modifying agents and the inhibition of Plasmodium falciparum growth: structure-activity relationships for betulinic acid analogues

The natural triterpene betulinic acid and its analogues (betulinic aldehyde, lupeol, betulin, methyl betulinate and betulinic acid amide) caused concentration-dependent alterations of erythrocyte membrane shape towards stomatocytes or echinocytes according to their hydrogen bonding properties. Thus, the analogues with a functional group having a capacity of donating a hydrogen bond (COOH, CH(2)OH, CONH(2)) caused formation of echinocytes, whereas those lacking this ability (CH(3), CHO, COOCH(3)) induced formation of stomatocytes. Both kinds of erythrocyte alterations were prohibitive with respect to Plasmodium falciparum invasion and growth; all compounds were inhibitory with IC(50) values in the range 7-28 microM, and the growth inhibition correlated well with the extent of membrane curvature changes assessed by transmission electron microscopy. Erythrocytes pre-loaded with betulinic acid or its analogues and extensively washed in order to remove excess of the chemicals could not serve as hosts for P. falciparum parasites. Betulinic acid and congeners can be responsible for in vitro antiplasmodial activity of plant extracts, as shown for Zataria multiflora Boiss. (Labiatae) and Zizyphus vulgaris Lam. (Rhamnaceae). The activity is evidently due to the incorporation of the compounds into the lipid bilayer of erythrocytes, and may be caused by modifications of cholesterol-rich membrane rafts, recently shown to play an important role in parasite vacuolization. The established link between erythrocyte membrane modifications and antiplasmodial activity may provide a novel target for potential antimalarial drugs.     

Investigation of Zinc Supplement Impact on the Serum Biochemical Parameters in Pulmonary Tuberculosis: A Double Blinded Placebo Control Trial

Background:Zinc (Zn) is nutritionally essential trace element, and thus deficiency may severely affect human health. The results of cross-sectional studies indicate that micronutrient deficiencies are common in patients with tuberculosis. Our goal is to investigate whether Zn supplementation can increase the effects of anti-TB treatment or not.Methods:Patients with newly diagnosed tuberculosis were divided in to 2 groups. One group (n= 37) received capsule contains 50 mg of elemental zinc (as zinc sulfate) for 6 months every other day (micronutrient group) and Group II (n= 37) received placebo. Both groups received the same anti-tuberculosis treatment recommended by the WHO. Clinical examination, BMI, chest X-ray, direct sputum examination, assessment of serum zinc levels (by atomic absorption spectrophotometry), and biochemical markers serum concentration (by using an RA1000 AutoAnalyzer) were carried out before and after 2- and 6-months anti-tuberculosis treatment.Results:Plasma zinc concentrations in the micronutrient group was higher than placebo group After treatment. In the placebo group increasing in SGOT and SGPT concentrations were significantly higher than micronutrient group after 2 months of treatment (p< 0.05). The significant changes (p< 0.05) were observed on the serum levels of total protein, albumin. Alkaline phosphatase (ALP) levels, serum creatinine, uric acid and urea in groups were not significantly different.Conclusion:Zinc supplementation results in earlier sputum smear conversion in the micronutrient group during the first 6 weeks. Increased body weight and serum zinc and serum albumin and decrease in total protein was observed in the micronutrient group.Key Words: Anti-tuberculosis treatment, Pulmonary tuberculosis, Zinc     

The Rapid Manufacture of Uniform Composite Multicellular-Biomaterial Micropellets,Their Assembly into Macroscopic Organized Tissues,and Potential Applications in Cartilage Tissue Engineering

We and others have published on the rapid manufacture of micropellet tissues, typically formed from 100–500 cells each. The micropellet geometry enhances cellular biological properties, and in many cases the micropellets can subsequently be utilized as building blocks to assemble complex macrotissues. Generally, micropellets are formed from cells alone, however when replicating matrix-rich tissues such as cartilage it would be ideal if matrix or biomaterials supplements could be incorporated directly into the micropellet during the manufacturing process. Herein we describe a method to efficiently incorporate donor cartilage matrix into tissue engineered cartilage micropellets. We lyophilized bovine cartilage matrix, and then shattered it into microscopic pieces having average dimensions < 10 μm diameter; we termed this microscopic donor matrix “cartilage dust (CD)”. Using a microwell platform, we show that ~0.83 μg CD can be rapidly and efficiently incorporated into single multicellular aggregates formed from 180 bone marrow mesenchymal stem/stromal cells (MSC) each. The microwell platform enabled the rapid manufacture of thousands of replica composite micropellets, with each micropellet having a material/CD core and a cellular surface. This micropellet organization enabled the rapid bulking up of the micropellet core matrix content, and left an adhesive cellular outer surface. This morphological organization enabled the ready assembly of the composite micropellets into macroscopic tissues. Generically, this is a versatile method that enables the rapid and uniform integration of biomaterials into multicellular micropellets that can then be used as tissue building blocks. In this study, the addition of CD resulted in an approximate 8-fold volume increase in the micropellets, with the donor matrix functioning to contribute to an increase in total cartilage matrix content. Composite micropellets were readily assembled into macroscopic cartilage tissues; the incorporation of CD enhanced tissue size and matrix content, but did not enhance chondrogenic gene expression.     

Tranilast enhances the anti-tumor effects of tamoxifen on human breast cancer cells in vitro

Background

Tamoxifen is the most widely used anti-estrogen for the treatment of breast cancer. Studies show that the combination therapy with other substances that helps the activity of tamoxifen. The objective of this study was to evaluate the effect of tamoxifen when used in combination with tranilast on human breast cancer cells.

Results

Two MCF-7 and MDA-MB-231 human breast cancer cell lines were treated with tamoxifen and/or tranilast. The cell viability and cytotoxicity was assessed using MTT and LDH assays; the apoptotic effects were examined by TUNEL assay, acridine orange/ethidium bromide staining and DNA laddering, also the expression levels of bax and bcl-2 genes were detected by real-time RT-PCR. The mRNA expression of TGF-β ligands and receptors examined using real-time RT-PCR and TGF-β1 protein secretion levels were also evaluated by ELISA assay. Inhibitory effect of these drugs on invasion and metastasis were tested by wound healing and matrigel invasion assay.We found that combination of these drugs led to a marked increase in growth and proliferation inhibition compared to either agent alone. Furthermore, bax and bcl-2 affected by tamoxifen and/or tranilast and resulted in a significant increase in bax and decrease in bcl-2 mRNA expression. In addition, treatment with tamoxifen and/or tranilast resulted in significant decreased in TGF-β1, 2, 3, TGF-βRI and II mRNA and TGF-β1 protein levels while TGF-βRIII mRNA level was increased and invasion was also inhibited.

Conclusions

These findings indicate that tranilast, by synergistic effect, enhances the activity of tamoxifen and the TGF-β pathway is a target for this combination therapy, therefore; we propose that this combined treatment may be suitable selection in prevention of breast cancer.