Pathology of tissue loss (white syndrome) in Acropora sp. corals from the Central Pacific

We performed histological examination of 69 samples of Acropora sp. manifesting different types of tissue loss (Acropora White Syndrome-AWS) from Hawaii, Johnston Atoll and American Samoa between 2002 and 2006. Gross lesions of tissue loss were observed and classified as diffuse acute, diffuse subacute, and focal to multifocal acute to subacute. Corals with acute tissue loss manifested microscopic evidence of necrosis sometimes associated with ciliates, helminths, fungi, algae, sponges, or cyanobacteria whereas those with subacute tissue loss manifested mainly wound repair. Gross lesions of AWS have multiple different changes at the microscopic level some of which involve various microorganisms and metazoa. Elucidating this disease will require, among other things, monitoring lesions over time to determine the pathogenesis of AWS and the potential role of tissue-associated microorganisms in the genesis of tissue loss. Attempts to experimentally induce AWS should include microscopic examination of tissues to ensure that potentially causative microorganisms associated with gross lesion are not overlooked.     

Mathematical treatment of water movement in plant cells and tissue: a review

Abstract. Over the past three decades, many contributions have been made to the development of a mathematical basis for describing water transport in plant cells and tissue. This review paper attempts to summarize the more significant contributions and to outline the concepts upon which the various mathematical analyses are founded.
The paper itself is divided into three major sections. Section I deals with the quantitative water relations of single plant cells. Basic equations are developed which describe the water statics and water dynamics of such cells. Included is a discussion of the theory and methods for measuring the various parameters (permeabilities, cell wall elastic moduli, etc.) which enter into the development. The section closes with a presentation of circuit analog models for single plant cells.
Section II is devoted to a review and development of the water relations of plant tissues which contain numerous cells in series. Following a historical overview, various existing models are derived and physical tissue properties which enter the derivation are identified. The concept of 'local equilibrium' is discussed and circuit analog models for single cells are generalized and applied to several cells in series.
The final section contains two example applications of water transport theory as it applies to plant tissue. One application involves radial water movement in a soybean hypocotyl while the other deals with water transport in a growing root tip. A summary at the end of the section is largely devoted to a discussion of the limitations of mathematical models dial are presently available.     

Experimental transmission and tissue tropism of Macrobrachium rosenbergii nodavirus (MrNV) and its associated extra small virus (XSV)

White tail disease (WTD) was found to be a serious problem in hatcheries and nursery ponds of Macrobrachium rosenbergii in India. The causative organisms have been identified as M. rosenbergii nodavirus (MrNV) and its associated extra small virus (XSV). Experimentally transmitted to healthy animals, they caused 100% mortality in post-larvae but failed to cause mortality in adult prawns. The RT-PCR assay revealed the presence of both viruses in moribund post-larvae and in gill tissue, head muscle, stomach, intestine, heart, hemolymph, pleopods, ovaries and tail muscle, but not in eyestalks or the hepatopancreas of experimentally infected adult prawns. The presence of these viruses in ovarian tissue indicates the possibility of vertical transmission. Pleopods have been found to be a suitable organ for detecting these viruses in brooders using the RT-PCR technique.     

Pathology associated with Cryptobia infection in a summer flounder (Paralichthys dentatus)

A laboratory-held summer flounder (Paralichthys dentatus) became moribund and presented gross ulcerative and hemorrhagic lesions, concomitant with a space-occupying lesion in the abdominal cavity and a prolapsed rectum. Edema, hemorrhage, and necrosis of the intestine and edema of the stomach wall were noted upon post-mortem examination. Microscopic examination revealed large numbers of Cryptobia in the submucosa of the gut and in the liver.     

PARP-1 and its associated nucleases in DNA damage response

Poly(ADP-ribose) (PAR) polymerase-1 (PARP-1) acts as a DNA damage sensor. It recognizes DNA damage and facilitates DNA repair by recruiting DNA repair machinery to damage sites. Recent studies reported that PARP-1 also plays an important role in DNA replication by recognizing the unligated Okazaki fragments and controlling the speed of fork elongation. On the other hand, emerging evidence reveals that excessive activation of PARP-1 causes chromatin DNA fragmentation and triggers an intrinsic PARP-1-dependent cell death program designated parthanatos, which can be blocked by genetic deletion or pharmacological inhibition of PARP-1. Therefore, PARP-1 plays an essential role in maintaining genomic stability by either facilitating DNA repair/replication or triggering DNA fragmentation to kill cells. A group of structure-specific nucleases is crucial for executing DNA incision and fragmentation following PARP-1 activation. In this review, we will discuss how PARP-1 coordinates with its associated nucleases to maintain genomic integrity and control the decision of cell life and death.     

Metformin (dimethyl-biguanide) induced DNA damage in mammalian cells

Metformin (dimethyl-biguanide) is an insulin-sensitizing agent that lowers fasting plasma-insulin concentration, wherefore it's wide use for patients with a variety of insulin-resistant and prediabetic states, including impaired glucose tolerance. During pregnancy it is a further resource for reducing first-trimester pregnancy loss in women with the polycystic ovary syndrome. We tested metformin genotoxicity in cells of Chinese hamster ovary, CHO-K1 (chromosome aberrations; comet assays) and in mice (micronucleus assays). Concentrations of 114.4 μg/mL and 572 μg/mL were used in in vitro tests, and 95.4 mg/kg, 190.8 mg/kg and 333.9 mg/kg in assaying. Although the in vitro tests revealed no chromosome aberrations in metaphase cells, DNA damage was detected by comet assaying after 24 h of incubation at both concentrations. The frequency of DNA damage was higher at concentrations of 114.4 μg/mL. Furthermore, although mortality was not observed in in vitro tests, the highest dose of metformin suppressed bone marrow cells. However, no statistically significant differences were noted in micronuclei frequencies between treatments. In vitro results indicate that chronic metformin exposure may be potentially genotoxic. Thus, pregnant woman undergoing treatment with metformin should be properly evaluated beforehand, as regards vulnerability to DNA damage.     

Gut associated lymphoid tissue in the cotton rat (Sigmodon hispidus) and its response to protein restriction.

We examined age and nutritional related changes in the distribution and size of gut associated lymphoid tissues in the intestinal tract of cotton rats (Sigmodon hispidus). Peyer's patches in the small intestine are prominent, ranging from four to 13, and increase in size (surface area) with age. The average Peyer's patch in the adult cotton rat measured 23.9 mm2. Lymphoid tissue in the cecum was primarily limited to a large aggregate located in the vermiform appendix, which increased in size with age. Age related changes in the number of visible lymphoid follicles in the large intestine were highly significant, increasing from 24.8 in juveniles to 45.1 in adults. Weights of dissectable Peyer's patch tissue in animals consuming a low protein diet were significantly lower in juveniles and greater in subadults compared to those on high protein diets. Relative weights of Peyer's patch tissue averaged 84 to 95% more in low protein-fed animals than in the group on the high quality protein diet. Our results suggest that peripheral lymphoid tissues in wild cotton rats are more resistant to protein deficiencies than other tissues in the body and could be a useful index for assessing nutritional status.     

Antigen retrieval in cryostat tissue sections and cultured cells by treatment with sodium dodecyl sulfate (SDS)

A simple method for antigen retrieval in tissue sections and cell cultures is described. Because many antibodies recognize denatured proteins on western blots, but are poorly reactive by immunocytochemistry, the effect of applying sodium dodecyl sulfate (SDS) to cryostat sections of tissues and to cell cultures prior to immunostaining was examined. In many cases, a 5-min pretreatment with 1% SDS produced a dramatic increase in staining intensity by indirect immunofluorescence. Among the antibodies tested that showed a positive effect of SDS were an anti-Na/K-ATPase monoclonal antibody, an anti-AE1/2 anion exchanger polyclonal antipeptide antibody, a monoclonal anti-caveolin antibody, and an anti-rab4 monoclonal antibody. In other cases, including antibodies against gp330, aquaporin 1, and aquaporin 2, no effect of SDS was detected. The results show that SDS treatment can be used as a simple method of antigen retrieval in cryostat sections and on cultured cells. In some cases, antigens were not detectable without pretreatment with SDS.     

DNA damage and repair in epithelial (mucous) cells and crypt cells from isolated colon

Colon epithelium is made up of two general classes of cells, surface cells which are post-mitotic and crypt cells which contain the proliferative population. Their relative vulnerability to environmental damage and ability to perform DNA repair are important issues in colon carcinogenesis. DNA damage and repair was studied by the nucleoid sedimentation method in freshly isolated crypt cells for comparison with previous studies of post-mitotic surface epithelial cells. Suspensions of crypt cells were isolated from preparations of mouse colon by a series of sequential incubations in buffer containing 1.5 mM EDTA. Treatment of crypt cells for 30 min with 1.2 X 10(-6) M methyl methane sulfonate (MMS), photoaffinity labeling with 1 X 10(-6) M ethidium monoazide, lithocholic acid (2.5 X 10(-4) M) treatment for 1 h or X-irradiation at 1500 rads resulted in single-strand breaks in the DNA, which were repaired after 2 h of additional incubation. Interestingly, X-rays at 1000 rads and lithocholic acid (LA) (2.5 X 10(-6) M) after 30 min incubation failed to produce the detectable shift in nucleoid sedimentation characteristic of single-strand breaks, perhaps due to rapid repair by these proliferative cells. UV-irradiation failed to provoke strand incision as was also observed for the superficial post-mitotic cells in the previous studies. These studies showed the feasibility of studying DNA damage and repair processes in these two classes of colon epithelial cells in response to specific carcinogenic insult.     

The symbiotic water mite Unionicola formosa (Acari: Unionicolidae) ingests mucus and tissue of its molluscan host

Unionicola formosa is a symbiotic water mite that passes most of its life cycle in the mantle cavity of freshwater mussels. Although mites of this genus are often referred to as parasitic, little is known about their nutritional biology. A few species reportedly pierce the gill of a host mussel and ingest tissue or hemolymph. The present study was undertaken to identify possible sources of nutrition for U. formosa. To determine if mites ingested particulate matter in the mucous strand produced by a mussel during feeding, mussels with resident mites were exposed to a suspension of fluorescent microspheres. There was no evidence that U. formosa ingested the beads. Histochemical staining did, however, indicate a mucous material present in the midgut of the mites. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic assays revealed a high molecular weight component, consistent with a mucopolysaccharide, present both in the mussel gill and the mites. Results from western blots and an immunoaffinity binding assay with antibodies against mussel gill tissue and hemolymph also indicated that mites ingested host tissue. Whereas U. formosa probably does not ingest particulate material acquired by its host's suspension feeding, it is apparent that this mite utilizes host mucus, gill tissue, or hemolymph for at least part of its nutrition.     

Iron-induced oxidative damage in colon carcinoma (Caco-2) cells

Intestinal epithelial cells have an active apical iron uptake system that is involved in the regulated absorption of iron. By the action of this system, intestinal cells acquire increasing amounts of iron with time. Since intracellular reactive iron is a source of free radicals and a possible cause of colon carcinoma, this study analyzed the oxidative damages generated by iron accumulation in Caco-2 cells. Cells cultured with increasing concentrations of iron increased both total intracellular iron and the reactive iron pool, despite an active IRE/IRP system, which regulates intracellular iron levels. Increasing concentrations of iron resulted in increased protein oxidative damage, as shown by the immunoreactivity for 4-hydroxy-2-nonenal-modified proteins, and markedly induced DNA oxidation determined by 8-hydroxy-2'-deoxyguanidine production. Iron also impaired cell viability, resulting in increased cell death after 6 days of culture. In summary, iron accumulation by intestinal Caco-2 cells correlated with oxidative damage to proteins and DNA. Oxidative damage finally resulted in loss of cell viability. The Fe-induced oxidative damage observed may be relevant in understanding the cascade of events associated with iron-mediated colon carcinogenesis.     

Poly(ADP-ribose) metabolism in X-irradiated Chinese hamster cells: its relation to repair of potentially lethal damage

Nicotinamide-adenine dinucleotide (NAD+) is the substrate used by cells in poly(ADP-ribose) synthesis. X-irradiation of log-phase Chinese hamster cells caused a rapid decrease in NAD+ levels which was linearly dependent on radiation dose. The activity of ADP-ribosyl transferase ( ADPRT ) also increased linearly with radiation dose. The decrease of NAD+ was slower, and the increase in ADPRT activity was less pronounced, in a radiation sensitive line, V79- AL162 /S-10. An inhibitor of ADPRT , m-aminobenzamide, largely prevented the depletion of cellular NAD+ and reduced the rate at which ADPRT activity disappeared during post-irradiation incubation. Post-irradiation treatment with hypertonic buffer or with medium containing D2O--which inhibit repair of radiation-induced potentially lethal damage--enhanced the depletion of NAD+ and prevented the reduction in ADPRT activity following irradiation. The characteristics of the effects of treatment with hypertonic buffer on NAD+ metabolism were qualitatively similar to the effects that such treatment has on radiation-induced cell killing. These results suggest that poly(ADP-ribose) synthesis after irradiation plays a role in the repair of potentially lethal damage.     

P relaxation responses associated with n(2)/o(2) diffusion in soybean nodule cortical cells and excised cortical tissue

N₂-fixing Bradyrhizobium japonicum nodules and cortical tissue derived from these nodules were examined in vivo by ³¹P nuclear magnetic resonance (NMR) spectroscopy. Perfusion of the viable nodules and excised cortical tissue with O₂ followed by N₂ or Ar caused a loss of orthophosphate (Pi) resonance magnetization associated with the major portion of acidic Pi (δ 0.9 ppm, pH 5.5) residing in the cortical cells. Resumption of O₂ perfusion restored approximately 80% of the intensity of this peak. Detailed examination of the nuclear relaxation processes, spin-lattice relaxation time (T₁), and spin-spin relaxation time (T₂), under perfusion with N₂ or Ar as opposed to O₂, indicated that loss of signal was due to T₁ saturation of the acidic Pi signal under the rapid-pulsed NMR recycling conditions. In excised cortical tissue, Pi T₁, values derived from biexponential relaxation processes under perfusing O₂ were 59% 3.72 ± 0.93 s and 41% 0.2 ± 0.08 s, whereas under N₂ these values were 85% 7.07 ± 1.36 s and 15% 0.39 ± 0.07 s. The T₁ relaxation behavior of whole nodule vacuolar Pi showed the same trend, but the overall values were somewhat shorter. T₂ values for cortical tissue were also biexponential but were essentially the same under O₂ (38% 0.066 ± 0.01 s and 63% 0.41 ± 0.08 s) and N₂ (39% 0.07 ± 0.01 s and 61% 0.37 ± 0.01 s) perfusion. Soybean (Glycine max) root tissue as well as Pi solutions exhibited single exponential T₁ decay values that were not altered by changes in the perfusing gas. These data indicate that oxygen induces a change in the physical environment of phosphate in the cortical cell tissue. Although under certain conditions oxygen has been observed to act as a paramagnetic relaxation agent, model T₁ experiments demonstrate that O₂ does not significantly influence Pi relaxation in this manner. Alternatively, we suggest that an increase in solution viscosity brought on by the production of an occlusion glycoprotein (under O₂ perfusion) is responsible for the observed relaxation changes.     

Hydroxyl radicals (*OH) are associated with titanium dioxide (TiO(2)) nanoparticle-induced cytotoxicity and oxidative DNA damage in fish cells

TiO(2) nanoparticles (< 100 nm diameter) have been reported to cause oxidative stress related effects, including inflammation, cytotoxicity and genomic instability, either alone or in the presence of UVA irradiation in mammalian studies. Despite the fact that the aquatic environment is often the ultimate recipient of all contaminants there is a paucity of data pertaining to the potential detrimental effects of nanoparticles on aquatic organisms. Therefore, these investigations aimed to evaluate the potential cytotoxic and genotoxic effects of TiO(2) nanoparticles on goldfish skin cells (GFSk-S1), either alone or in combination with UVA. Whilst neutral red retention (NRR) assay (a measure of lysosomal membrane integrity) was used to evaluate cell viability, a modified Comet assay using bacterial lesion-specific repair endonucleases (Endo-III, Fpg) was employed to specifically target oxidative DNA damage. Additionally, electron spin resonance (ESR) studies with different spin traps were carried out for qualitative analysis of free radical generation. For cell viability, TiO(2) alone (0.1-1000 microg ml(-1)) had little effect whereas co-exposure with UVA (0.5-2.0 kJm(-2)) caused a significant dose-dependent decrease which was dependent on both the concentration of TiO(2) and the dose of UVA administered. For the Comet assay, doses of 1, 10 and 100 microg ml(-1) in the absence of UVA caused elevated levels of Fpg-sensitive sites, indicating the oxidation of purine DNA bases (i.e. guanine) by TiO(2). UVA irradiation of TiO(2)-treated cells caused further increases in DNA damage. ESR studies revealed that the observed toxic effects of nanoparticulate TiO(2) were most likely due to hydroxyl radical (OH) formation.