OBJECTIVE: To determine in situ using TEM the balance of apoptosis and necrosis in the articular cartilage of patients with inflammatory (rheumatoid arthritis and seronegative spondyloarthritis) and degenerative (osteoarthritis) joint diseases and to establish possible correlation between the cell death rate and the matrix vesicles formation.
METHODS: Cartilage samples of the knee joint were obtained from patients with rheumatoid arthritis (RA, 18 cases), osteoarthritis (OA, 22 cases), Reiter's disease (RD, 9 cases), peripheral form of the ankylosing spondyloarthritis (AS, 6 cases) and psoriatic arthritis (PA, 6 cases) during arthroscopy or knee surgery. Normal samples taken from autopsy cases without a history of joint diseases were used as control. Samples were processed for TEM with subsequent semi-quantitative estimation of the cell death rate in the superficial, middle and deep zone of non-calcified articular cartilage, and computer-aided ultramorphometric evaluation of the matrix vesicles of different types.
RESULTS: Both apoptotic and necrotic cell death could be identified in the cartilage of patients with inflammatory joint diseases, including seronegative spondyloarthritides and degenerative arthropathies. Apoptosis dominated over necrosis in all examined arthritides, including RA patients in which necrosis of the chondrocyte was the most frequent among arthropathies, while the highest apoptotic cell death rate was discovered in OA in which it correlated with the volume and numeric density of the matrix vesicles. These data provide evidence that apoptosis may contribute to the cartilage breakdown not only in RA and OA but also in the seronegative spondyloarthritides, which had a significantly higher apoptotic rate than the normal cartilage.
Scanning Electron Microscopy (SEM) and Transmission Electron Microscopy (TEM) evaluation were carried out in the in vivo skin construct using fibrin as biomaterial. To investigate its progressive remodeling, nude mice were grafted and the Extracellular Matrix (ECM) components were studied at four and eight weeks post-grafting. It was discovered that by 4 weeks of remodeling the skin construct acquired its native structure.
Bone is unique in its ability to adapt structure to functional requirements, but as is all too obvious in an ever-ageing population it is susceptible to a number of degenerative diseases. Therefore there is an increasing need for materials for bone replacement. Clearly, the ideal material with which to replace bone, would be bone itself, but the major problem now facing us is that there is an insufficient supply of the natural bone to satisfy the clinical requirements. Hence, there is a need for the development of chemically synthesised bone graft substitutes
OBJECTIVE: To evaluate the range of activation changes of polymorphonuclear leukocytes (PMN) and the ratio of apoptosis and necrosis in synovial effusions of patients with various arthropathies, and to reveal possible correlations with clinical variants of joint inflammation.
METHODS: Synovial effusions were aspirated from the knee joints of patients with rheumatoid arthritis (RA, 28 cases), and seronegative spondyloarthritides (SSA): Reiter's disease (RD, 9 cases), peripheral form of the ankylosing spondyloarthritis (6 cases) and psoriatic arthritis (6 cases); and primary osteoarthritis (OA, 9 cases). Cytospin preparations were processed for transmission electron microscopy and assessed for the incidence of apoptosis, necrosis, and cytophagocytic cells (CPC) in the synovial fluid (SF). The range of activation changes of the neutrophil granulocytes, the dominating cell population in the arthritic SF, was evaluated.
RESULTS: In all arthropathies under investigation most of the synovial effusion cells had intact ultrastructure with a certain amount of apoptotic cells dominating over the cells with signs of necrosis, and a few CPC. The highest rate of apoptosis was discovered in the synovial effusions of patients with RA, the lowest in those with OA, while the rate of CPC among the inflammatory joint diseases was the lowest in RA. In RA the current disease activity correlated with the incidence of apoptotic cells and CPC, while the clinical stage was related only to the CPC rate. These data suggest that in RA, despite exposure to the anti-apoptotic signals, apoptosis of the synovial effusion PMN is maintained at a significantly higher level than in non-rheumatoid arthropathies, both inflammatory (SSA) and degenerative (OA), providing elimination of the neutrophils accumulating in the joint cavity and thus stimulating resolution of the joint inflammation.
Currently, breast cancer is the leading cause of cancer-related death in women. Therefore, there is an urgent need to develop alternative therapeutic measures against this deadly disease. Here, we report the cytotoxicity activity and the mechanism of cell death exhibited by the methanol extract prepared from Pereskia bleo (Kunth) DC. (Cactaceae) plant against human breast carcinoma cell line, T-47D. In vitro cytotoxicity screening of methanol extract of Pereskia bleo plant indicated the presence of cytotoxicity activity of the extract against T-47D cells with EC50 of 2.0 microg/ml. T-47D cell death elicited by the extract was found to be apoptotic in nature based a clear indication of DNA fragmentation which is a hallmark of apoptosis. In addition, ultrastructural analysis also revealed apoptotic characteristics (the presence of chromatin margination and apoptotic bodies) in the extract-treated cells. RT-PCR analysis showed the mRNA expression levels of c-myc, and caspase 3 were markedly increased in the cells treated with the plant extract. However, p53 expression was only slightly increased as compared to caspase 3 and c-myc. Thus, the results from this study strongly suggest that the methanol extract of Pereskia bleo may contain bioactive compound(s) that caused breast carcinoma, T-47D cell death by apoptosis mechanism via the activation of caspase-3 and c-myc pathways.
Leydig and Sertoli cells of the immature lesser mouse deer testes, obtained in East Malaysia, were observed using light and transmission electron microscopy (TEM). The testes were fixed in 5% glutaraldehyde, post-fixed in 1% OsO4, dehydrated in ethanol, and embedded in Araldite M. Serial semi-thin sections were cut, stained with toluidine blue and observed using light microscopy. Serial ultra-thin sections were cut, stained with uranyl acetate and lead citrate, and examined using TEM. As a result, ultrastructurally, two types of underdeveloped filament bundles were infrequently recognized in Leydig cells, but not in other testicular cells. One type was the underdeveloped bundles of actin filaments (approximately 5 nm in diameter), which were found in the nucleus of Leydig cells. The other type was the underdeveloped bundles of intermediate filaments (approximately 10 nm in diameter), which were found in the cytoplasm of Leydig cells. A multivesicular nuclear body (MNB)--specifically present in the Sertoli cell nucleus of ruminant testes--was infrequently observed. The MNB is situated in the vicinity of nuclear membrane, still in an underdeveloped stage.
Three strains of Spirulina platensis isolated from different locations showed capability of synthesizing poly(3-hydroxybutyrate) [P(3HB)] under nitrogen-starved conditions with a maximum accumulation of up to 10 wt.% of the cell dry weight (CDW) under mixotrophic culture conditions. Intracellular degradation (mobilization) of P(3HB) granules by S. platensis was initiated by the restoration of nitrogen source. This mobilization process was affected by both illumination and culture pH. The mobilization of P(3HB) was better under illumination (80% degradation) than in dark conditions (40% degradation) over a period of 4 days. Alkaline conditions (pH 10-11) were optimal for both biosynthesis and mobilization of P(3HB) at which 90% of the accumulated P(3HB) was mobilized. Transmission electron microscopy (TEM) revealed that the mobilization of P(3HB) involved changes in granule quantity and morphology. The P(3HB) granules became irregular in shape and the boundary region was less defined. In contrast to bacteria, in S. platensis the intracellular mobilization of P(3HB) seems to be faster than the biosynthesis process. This is because in cyanobacteria chlorosis delays the P(3HB) accumulation process.
Sexually mature males of Bactrocera papayae are strongly attracted to and consume methyl eugenol (ME). Upon consumption, ME is biotransformed to two phenylpropanoids, 2-allyl-4,5-dimethoxyphenol (DMP) and (E)-coniferyl alcohol (CF), that are transported in the hemolymph, sequestered and stored in the rectal glands, and subsequently released as sex and aggregation pheromones during courtship. To date, very little work on the ultrastructure and anatomy of the rectal gland has been done, and the accumulation of phenylpropanoids in the rectal glands of males has not been observed visually. Our objectives are to describe the anatomy and fine structures of the rectal glands of males and females and to observe the accumulation of autofluorescent compounds in the rectal glands of males. The rectal glands of males and females have four rectal papillae with each papilla attached to a rectal pad. The rectal pads protrude from the rectal gland as the only surfaces of the gland that are not surrounded by muscles. The rectal papillae of ME-fed males had oil droplets and autofluorescent compounds that were absent from those of ME-deprived males. The autofluorescent compounds accumulated in the rectal sac, which is an evagination that is not found in rectal glands of females. The accumulation of these compounds increased with time and reached maximum at a day post-ME feeding and decreased thereafter. This trend is similar to the accumulation pattern of phenylpropanoids, CF and DMP in the rectal gland.
Palm kernel oil, palm olein, crude palm oil and palm acid oil were used for the synthesis of poly (3-hydroxybutyrate-co-3-hydroxyhexanoate) [P(3HB-co-3HHx)] by a mutant strain of Wautersia eutropha (formerly Ralstonia eutropha) harboring the Aeromonas caviae polyhydroxyalkanoate (PHA) synthase gene. Palm kernel oil was an excellent carbon source for the production of cell biomass and P(3HB-co-3HHx). About 87% (w/w) of the cell dry weight as P(3HB-co-3HHx) was obtained using 5 g palm kernel oil/l. Gravimetric and microscopic analyses further confirmed the high PHA content in the recombinant cells. The molar fraction of 3HHx remained constant at 5 mol % regardless of the type and concentration of palm oil products used. The small amount of 3HHx units was confirmed by 13C NMR analysis. The number average molecular weight (M(n)) of the PHA copolymer produced from the various palm oil products ranged from 27 0000 to 46 0000 Da. The polydispersity was in the range of 2.6-3.9.
A study of the anterior adhesive apparatus (head organs) of Bravohollisia gussevi Lim, 1995 was carried out using light and electron microscopy. The anterior adhesive apparatus or head organs in B. gussevi comprise 6 circular openings or apertures in the antero-lateral region, associated pits lined with specialized microvillous tegument that differ from the general body tegument, a bundle of ducts, and uninucleate gland cells located lateral to the pharynx. The uninucleate glands of the anterior adhesive apparatus (head organs) comprise 2 types of cells, one kind of cell producing rod-like bodies (S1) and the other oval bodies (S2). The S1 bodies are filled with numerous, less electron-dense vesicles in an electron-dense matrix, while S2 bodies have no vesicles but contain a more homogeneous electron-dense matrix. Interlinking band-like structures were observed between S1 bodies. Similar band-like structures were found between S2 bodies. The formation of S1 bodies was followed by transmission electron microscopy. However, the formation of S2 bodies was unclear and could not be resolved. Uniciliated structures were also observed around the openings of the anterior adhesive apparatus. Each uniciliated structure is usually associated with an opening of a gland cell producing granular, electron-dense, secretory bodies, which differ from the secretions produced by the lateral gland cells of the anterior adhesive apparatus.
Transmission electron microscopy (TEM) can provide high resolution imaging of biological specimens. The study is to establish the effects of a modified glutaraldehyde (GA) compare to the standard GA fixation on Acanthamoeba castellanii from TEM perspectives and thus provide precise and accurate information on the ultrastructure studies of the parasite. By increasing the contrast, the ultrastructures of the parasite were more evident. The TEM images were obtained from parasites fixed with the modified GA and the standard GA and then the area of the nucleus and the gray values of the image of the nucleus of the parasites were measured. The mean areas of the nucleus were found to be significantly reduced in the standard GA fixed parasites (12210.4 nm2) compared to the modified GA fixed parasites (8676.3 nm2) (p < 0.05). The mean gray values of the image were significantly reduced from 2024 in the standard GA fixed parasites (2024) to the modified GA fixed parasites (1636) (p < 0.05). The study shows that the modified GA produced significantly better contrast on TEM images of the A. castellanii compared to the standard GA. This was because the modified GA generated more free water molecules during fixation and the uptake of modified GA by the nucleus of the parasite organizing all protein constituents in the cell into a more closely packed configuration than that of the standard GA. With such properties, the modified GA is a better fixative providing better images for ultrastructures of the parasite.
The Brassica napus secondary embryogenesis system requires no exogenous growth regulator to stimulate embryo development. It is stable embryogenically over a long period of culture and has a distinct pre-embryogenic stage. This system was used to investigate the morphological and cellular changes occurring in the embryogenic tissue compared to non-embryogenic tissue using various microscopy techniques. A unique ultrastructural feature designated the extracellular matrix (ECM) was observed on the surface of pre-embryogenic embryoids but not on the non-embryogenic individuals. The ECM layer was found to be dominant in the pre-embryogenic stage and reduced to fragments during embryo growth and development in mature embryogenic tissue. This is a novel aspect of the phenotype previously unreported in the Brassica system. This structure might be linked to acquisition of embryogenic competence.
The amoeboid form of Blastocystis hominis has been reported infrequently, and its morphological descriptions have yielded conflicting and confusing reports. In the present study, we used the amoeboid forms seen predominantly in symptomatic patients infected with Blastocystis to provide detailed descriptions on the fine surface structure and intracellular morphology. Scanning electron microscopy revealed the irregular shape of the amoeboid form, with an intercalated fibrillar structure and a highly convoluted surface with deep indentations and projected pseudopodia. Transmission electron microscopy showed the existence of two types of amoeboid forms of B. hominis in in vitro culture, one with a large central vacuole containing tiny electron-dense particles while the other contains multiple small vacuoles in the cytoplasm. A surface coat with varying thickness surrounded the amoeboid form, which also showed prominent, extended pseudopodia of varying shape. Irregularly shaped mitochondrion-like organelles with prominent cristae, lipid inclusions, and multiple vacuoles were frequently seen in close proximity with the pseudopodia. The characteristic nucleus with a crescentic band of electron-dense chromatin material was also seen.
The authors describe the case of a 50-year-old man with chronic progressive external ophthalmoplegia (CPEO), diabetes mellitus (DM), and coronary artery disease. The patient had no cardiac conduction abnormalities. During coronary artery bypass surgery, his heart and two skeletal muscles were biopsied. All three muscles showed ragged red fibers. The heart muscle showed significant glycogen accumulation. Analysis of mitochondrial DNA (mtDNA) showed a 5019-base-pair deletion, with no duplications. There were morphologically abnormal mitochondria in all 3 muscles, with clinically apparent difference in preservation of function. The combination of diabetes mellitus and mtDNA deletion is fortuitous, as they can be causally linked. The cardiac pathology allows speculation about the possible adaptive processes that may occur in the heart in DM. There are few reported cases with CPEO and excess glycogen in the heart. Most show deposition of fat and poorer clinical outcomes as compared to those with glycogen deposition. This observation may lend support to the hypothesis that in the myocardium, adaptive responses are mediated via changes in glucose handling, whereas alterations in fat metabolism likely represent maladaptation.
Cytomegalovirus (CMV) congenital infection is the major viral cause of well-documented birth defects in human. Because CMV is species-specific, the main obstacle to developing animal models for congenital infection is the difference in placental architecture, which preludes virus transmission across the placenta. The rat placenta, resembling histologically to that of human, could therefore facilitate the study of CMV congenital infection in human.
Ice cream mixes containing 33.4% total solids including 10% fat, 11.1% milk solid-non fat (MSNF), 12% sugar, 0.35% commercial blend of emulsifier/ stabiliser and water were produced. The blending of PO with AMF were conducted at three different ratios 30: 70, 50: 50 and 70: 30, respectively. The experimental ice cream mixes were compared with a control ice cream mix prepared from AMF. The flow properties were measured after ageing at 0, 1, 1.5, 2 and 24 h and determined using a controlled stress rheometer (Haake RS 100). The Power Law and Casson equation was employed to estimate the yield stress of an ice cream mixes. The regression coefficients (r) was represented well by the Casson model (r > 0.99) for all the samples, indicating goodness of fit. The profiles of the consistency coefficients (K(c)) were quite similar for all experimental samples, which could be attributed to the fact that all the samples exhibited similar viscoelastic behaviour. The flow behaviour index (n) of an ice cream mix prepared from PO and their blends with AMF were less then 1.0 (range 0.04-0.08) indicating that they were psuedoplastic fluid. The eta(o) at shear rate 20(-1) indicated higher degree of viscosity in AMF.
Iron and 4-(methylamino)benzoic acid have been successfully incorporated into silica extracted from rice husk. The silica/Fe/amine complex, RH-Fe(5% amine), showed a ca. 24% increase in specific surface area compared to RH-Fe. This increase was attributed to the templated formation of regular pores. The XRD showed the RH-Fe(5% amine) to be amorphous. The Friedel-Crafts benzylation reaction with toluene using RH-Fe(5% amine) showed a drastic reduction in the di-substituted products to ca. 1.0%.
Blastocystis hominis has been regarded as an enigmatic parasite as many aspects of its basic biology remain uncertain. Many reproductive processes have been suggested for the organism; however, to date, only the binary fission has been proven. Plasmotomy is one of the modes of reproduction previously suggested to be seen in in vitro cultures. The present study provides trichrome and acridine orange staining evidence for the existence of nucleic acid suggestive of division of nucleus into multinucleate forms with the respective cytoplasm dividing giving rise to two or three progeny B. hominis. Transmission electron micrographs further confirmed that these daughter cells had respective surrounding surface coat, mitochondria, and vacuoles.
Alumina powder was synthesized from an aluminum precursor and studied using small angle neutron scattering (SANS) technique and complemented with transmission electron microscope (TEM). XRD measurement confirmed that the alumina produced was of high purity and highly crystalline D-phase. SANS examination indicates the formation of mass fractals microstructures with fractal dimension of about 2.8 on the alumina powder.