Suppressive activity of protease inhibitors from buckwheat seeds against human T-acute lymphoblastic leukemia cell lines

The buckwheat protease inhibitor designated BWI-1, a member of the potato inhibitor I family, inhibits trypsin, chymotrypsin, and subtilisin, whereas the buckwheat protease inhibitor designated BWI-2a, a novel protease inhibitor homologous to the vicilin family, inhibits only trypsin. We examined the suppressive activity of BWI-1 and BWI-2a against T-acute lymphoblastic leukemia (T-ALL) cells, such as JURKAT and CCRF-CEM, and human normal blood lymphocytes. Both inhibitors significantly suppressed the growth of T-ALL cells as judged by the soluble 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (tetrazolium/formazan assay). JURKAT cells showed slightly higher susceptibility to buckwheat inhibitors than CCRF-CEM cells. Modification of Arg residue(s) in inhibitors by 1,2-cyclohexandione inactivated their trypsin inhibitory activity, considerably abolishing their suppressive activity. This suggests that the trypsin inhibitory activity is involved in the suppression of growth of human T-ALL cell lines. It was further found that both inhibitors triggered programmed cell death (apoptosis) of these cell strains with DNA fragmentation.     

Essential oil of <Emphasis Type="Italic">Laurus nobilis</Emphasis> from Montenegro

Steam distilled oil from the shoots, separated leaves, and stem, as well as from the flower of laurel (Laurus nobilis), grown in Montenegro, were analyzed by GC and GC/MS. The yield of essential oil was as follow: 1.4% in young shoots, 1.5% in the separated leaves, and 0.7% in separated stems. The main constituents of all investigated oils were 1,8-cineole, methyleugenol, and α-terpinyl acetate. Besides, α-pinene, β-pinene, sabinene, and linalool were also present. It was interesting and important for commercial samples of laurel essential oil that there was no significant difference among the essential oil obtained from young shoots and those obtained from leaves and stem. The main constituents of the flower oil were 1,8-cineole (15.7%), β-caryophyllene (9.5%), γ-muurolene (7.1%), α-terpinyl acetate (6.5%), and methyleugenol (3.9%). Published in Khimiya Prirodnykh Soedinenii, No. 4, pp. 337–339, July–August, 2007.     

Design of a Novel Composite H2S‐Releasing Hydrogel for Cardiac Tissue Repair

The design of 3D scaffolds is a crucial step in the field of regenerative medicine. Scaffolds should be degradable and bioresorbable as well as display good porosity, interconnecting pores, and topographic features; these properties favour tissue integration and vascularization. These requirements could be fulfilled by hybrid hydrogels using a combination of natural and synthetic components. Here, the mechanical and biological properties of a polyethylene glycol‐fibrinogen hydrogel (PFHy) are improved in order to favour the proliferation and differentiation of human Sca‐1^pos cardiac progenitor cells (hCPCs). PFHys are modified by embedding air‐ or perfluorohexane‐filled bovine serum albumin microbubbles (MBs) and characterized. Changes in cell morphology are observed in MBs–PFHys, suggesting that MBs could enhance the formation of bundles of cells and influence the direction of the spindle growth. The properties of MBs as carriers of active macromolecules are also exploited. For the first time, enzyme‐coated MBs have been used as systems for the production of hydrogen sulfide (H₂S)‐releasing scaffolds. Novel H₂S‐releasing PFHys are produced, which are able to improve the growth of hCPCs. This novel 3D cell–scaffold system will allow the assessment of the effects of H₂S on the cardiac muscle regeneration with its potential applications in tissue repair.

     

Poly(l‐Lactic Acid)/Gelatin Fibrous Scaffold Loaded with Simvastatin/Beta‐Cyclodextrin‐Modified Hydroxyapatite Inclusion Complex for Bone Tissue Regeneration

Recently, the application of nanostructured materials in the field of tissue engineering has garnered attention to mediate treatment and regeneration of bone defects. In this study, poly(l ‐lactic acid) (PLLA)/gelatin (PG) fibrous scaffolds are fabricated and β‐cyclodextrin (βCD) grafted nano‐hydroxyapatite (HAp) is coated onto the fibrous scaffold surface via an interaction between βCD and adamantane. Simvastatin (SIM), which is known to promote osteoblast viability and differentiation, is loaded into the remaining βCD. The specimen morphologies are characterized by scanning electron microscopy. The release profile of SIM from the drug loaded scaffold is also evaluated. In vitro proliferation and osteogenic differentiation of human adipose derived stem cells on SIM/HAp coated PG composite scaffolds is characterized by alkaline phosphatase (ALP) activity, mineralization (Alizarin Red S staining), and real time Polymerase chain reaction (PCR). The scaffolds are then implanted into rabbit calvarial defects and analyzed by microcomputed tomography for bone formation after four and eight weeks. These results demonstrate that SIM loaded PLLA/gelatin/HAp‐(βCD) scaffolds promote significantly higher ALP activity, mineralization, osteogenic gene expression, and bone regeneration than control scaffolds. This suggests the potential application of this material toward bone tissue engineering.

     

Regenerative medicine and stem cell based drug discovery

As William Shakespeare beautifully described, increasing age often causes loss of tissue and organ function. The increase in average life expectancy in many countries is generating an aging society and an increase in age-related health problems. Regenerative medicine is expected to be a powerful actor in this drama, and stem cell technology may hold the key to the development of innovative treatments for acute and chronic degenerative conditions. This Review surveys the present situation and some future prospects for regenerative medicine and stem cell based drug discovery.     

Recruitment of endogenous stem cells for tissue repair

The traditional concept of stem cell therapy envisions the isolation of stem cells from patients, propagation and differentiation in vitro, and subsequent re-injection of autologous cells into the patient. There are many problems associated with this paradigm, particularly during the in vitro manipulation process and the delivery and local retention of re-injected cells. An alternative paradigm that could be easier, safer, and more efficient, would involve attracting endogenous stem cells and precursor cells to the defect site for new tissue regeneration. Hepatocyte growth factor (HGF), a pleiotropic cytokine of mesenchymal origin, exerts a strong chemoattractive effect on mesenchymal stem cells (MSCs) and neural stem cells (NSCs), and induces migration of MSCs in vitro. However, HGF undergoes rapid proteolysis in vivo, which results in a very short lifetime of the bioactive cytokine. To maintain the therapeutic level of HGF at the defect site necessary for endogenous stem cell recruitment, sustained, long-term, and localized delivery of HGF is required. Thiol-modified glycosaminoglycans hyaluronan (HA) and heparin (HP), combined with modified gelatin (Gtn), have been crosslinked with poly(ethylene glycol) diacrylate (PEGDA) to afford semisynthetic ECM-like (sECM) hydrogels that can both provide controlled growth factor release and permit cell infiltration and proliferation. Herein we compare the use of different sECM compositions for controlled release of HGF and concomitant recruitment of human bone marrow MSCs into the scaffold in vitro. [Figure: see text].     

Effect of fatty acid methyl esters from plastrum testudinis on proliferation of rat bone mesenchymal stem cells

The ointment of plastrum testudinis was extracted using petroleum ether, ether and dichloromethane sequentially and the extracts were methyl-esterified. The effects on the proliferation of bone marrow mesenchymal stem cells (bMMSCs) were examined by MTT[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay and flow cytometry analysis. The volatile components of the samples were studied by gas chromatography-mass spectrometry (GC-MS) and high performance liquid chromatography (HPLC). The results show that the methyl-esterified parts can promote the proliferation of stem cells and they all contain palmitic acid methyl ester. Palmitic acid methyl ester can promote proliferation when the concentration was 0.15 μg/μL. It may be concluded that the palmitic acid methyl ester is important for the methyl-esterified parts that have effects on proliferation. __________ Translated from Chinese Journal of Analytical Chemistry, 2007, 35 (10): 1400–1404 [译自: 分析化学]