Altered cytokeratin expression and differentiation induction during neoplastic transformation of cultured rat liver cells by nickel subsulfide

Rat liver T51 B cells were maintained in the presence of low concentrations of Ni(H) derived from Ni₃S₂ for 3–I5 months in culture in order to monitor cytokeratin, differentiation, and transformation patterns. Nickel exposures caused irreversible, heritable juxtanuclear aggregates of cytokeratin CKSS, which increased in size and complexity with prolonged nickel exposure, eventually resembling Mallory bodies and expressing glutamyltransferase. Altered cytokeratin expression was accompanied by induction of differentiation, with markers of both bile ductular cells and hepatocytes, such as induction of cytokeratin polypeptides CK39 and CK49, cell morphology, and cytokeratin filament network changes, whereas control cultures similarly maintained for long periods in culture remained unchanged. Altered cytokeratin expression was also accompanied by acquisition of transformation markers—loss of density dependence, progression toward calcium independence, and (benign) growth in nude mice. Observed cytokeratin aberrations may be a factor in nickel carcinogenesis, in view of the known affinity of the metal for cellular structural proteins, especially keratin, which play a role in maintenance of cell behavior.Abbreviations CK cytokeratin - GGT gamma-glutamyltransferase - HCM standard calcium medium - LCM low calcium medium (Ca²⁺ = 0.02 mM) - MB mallory body - NI neoplastic index (ratio of growth of cells in LCM/HCM) - Ni ni(II) used as leachate from Ni₃S₂ - PBS phosphate-buffered saline - SB sodium butyrate     

肾动脉内注射缓激肽对麻醉家兔肾伟入神经放电的双相激活

The effect of intrarenal artery injection of bradykinin (BK, 5.0 micrograms/kg) on multi- and single-unit recordings of afferent renal nerve activity (ARNA) was examined in anesthetized 48 rabbits. The results obtained are as follows. (1) There were two phases of increase in ARNA following intrarenal BK. The early phase occurred immediately while the delayed phase made its appearance about 7 min later. The degree of increase in ARNA of the delayed phase induced by intrarenal BK was more prominent than that in the early phase. BP was actually unaltered following intrarenal BK. (2) By pretreatment with indomethacin (Indo, 5.0 mg/kg), the delayed phase of increase in ARNA induced by intrarenal BK was attenuated, while the early phase was not affected. (3) Pretreatment with L-NAME (30 mg/kg) led the delayed phase to be blocked completely while the early phase was partially decreased. From the above-mentioned observations, it is concluded that intrarenal BK induces a significant increase in ARNA in two phases. The early phase may be due to the direct action of BK and partially due to the NO action, while the delayed phase may be attributed to the action of released prostaglandin and NO as a result of intrarenal BK.     

Induced sensitization to nickel in guinea pigs immunized with mycobacteria by injection of purified protein derivative with nickel

Nickel has been reported to be one of the most common causes of allergic contact dermatitis. Despite the fact that nickel is a frequent sensitizer in humans, establishing animal models for nickel allergy has met with considerable difficulties. In clinical cases, allergic contact hypersensitivity to nickel develops much more readily in inflamed skin than normal skin. In this study, we tried to induce nickel sensitization when inflammation has been evoked in guinea pigs immunized with mycobacteria followed by co-administration of a mycobacterial component with nickel. We first examined the delayed-type hypersensitivity (DTH) reaction of mycobacterial components such as the cell wall, cell membrane, 70S ribosomal fraction, cytoplasm, tuberculin purified protein derivative (PPD), RNA and DNA from Mycobacterium bovis BCG in guinea pigs immunized with live M. bovis BCG or heat killed M. tuberculosis. When PPD was used, the hypersensitivity reaction was strongest. Next, we tested whether PPD with nickel could induce nickel sensitivity in guinea pigs immunized with mycobacteria. Strong sensitization to nickel was achieved by injecting PPD with nickel. However, if too large an amount of PPD or nickel salts was used, sensitization to nickel decreased. In this way, sensitization of nickel developed much more easily in guinea pigs immunized with mycobacteria by injection of an appropriate amount of nickel at the inflammation site induced by a suitable amount of PPD.     

Effects of circadian phase and melatonin injection on anxiety-like behavior in nocturnal and diurnal rodents

Animals show daily rhythms in most bodily functions, resulting from the integration of information from an endogenous circadian clock and external stimuli. These rhythms are adaptive and are expected to be related to activity patterns, i.e., to be opposite in diurnal and nocturnal species. Melatonin is secreted during the night in all mammalian species, regardless of their activity patterns. Consequently, in diurnal species the nocturnal secretion of melatonin is concurrent with the resting phase, whereas in nocturnal species it is related to an increase in activity. In this research, we examined in three diurnal and three nocturnal rodent species whether a daily rhythm in anxiety-like behavior exists; whether it differs between nocturnal and diurnal species; and how melatonin affects anxiety-like behavior in species with different activity patterns. Anxiety-like behavior levels were analyzed using the elevated plus-maze. We found a daily rhythm in anxiety-like behavior and a significant response to daytime melatonin administration in all three nocturnal species, which showed significantly lower levels of anxiety during the dark phase, and after melatonin administration. The diurnal species showed either an inverse pattern to that of the nocturnal species in anxiety-like behavior rhythm and in response to daytime melatonin injection, or no rhythm and, accordingly, no response to melatonin.     

Regulation of leucogenesis by extracellular ubiquitin in rodents after chemically induced inhibition

To study the influence of intraperitoneal injected extracellular ubiquitin on regeneration of leucopoiesis calculation of nuclear cell count in bone marrow (BM) and peripheral blood (PB) smears stained with azure-eosin was performed. In the first, control group of animals inhibition of haematopoiesis achieved by means of 100 mg/kg cyclophosphamide LD₅₀ 50–200 mg/kg injection. Bone marrow and peripheral blood samples from the first group of rats had been taken at 24, 48, 72, 96 and 168 h points after injection of cytostatic. Animals of the second, test group were injected by 200 μg/mL ubiquitin 72 h later after cytostatic injection. Our experiments revealed that ubiquitin makes corrections in regeneration of leucopoiesis and leads to normalisation of the process. Ubiquitin regulates stem cell activity, normalizes the release of functional cells into bloodstream, supposedly retains progenitor cells in zones of differentiation and maturation, and restores the nuclear cell ratio in PB and BM. We suppose that obtained results are important for elucidation of new pathways of ubiquitinylation and give us possibilities to find new therapeutics for regeneration of leucopoiesis that is very essential for treatment of radiated bone marrow and chemotherapeutic side effects in cancer patients.     

Estimation of the complex of hemorheological parameters in erythrocytosis

The complex of hemorheological parameters in erythrocytosis was studied. Typical changes in the factors responsible for blood fluidity were detected. A decreased efficiency of oxygen transport was found under these conditions. The pronounced hemorheological changes were shown to depend on the erythrocytosis intensity.     

高原红细胞增多症患者肠道菌群变化研究

目的探究高原红细胞增多症患者肠道菌群变化及相关意义。方法招募西藏那曲地区血红蛋白(hemoglobin, HB)含量高于210 g/L的高原红细胞增多症患者12人,对照组(170 g/LHB210 g/L)42人,采集粪便、血液等样本。从中选取高原红细胞增多症患者9人,对照组16人粪便样本提取DNA,进行16S rRNA测序分析。结果高原红细胞增多症患者的血红蛋白水平以及红细胞比容显著高于对照组。红细胞增多症患者的肠道菌群整体结构改变不明显,多样性没有显著差异。红细胞增多症患者的粪便菌群出现厚壁菌门中梭状芽胞杆菌BB60属、纺锤链杆菌属、厌氧棒杆菌属、霍氏真杆菌,变形菌门中假单胞菌属,拟杆菌门中副拟杆菌属丰度的显著增高。这些菌属改变经KEGG预测分析显示与代谢紊乱相关。斯皮尔曼分析显示多种菌群改变与血红蛋白水平以及红细胞比容正相关。结论高原红细胞增多症患者肠道菌群出现紊乱,菌群变化与代谢紊乱和红细胞增多相关。     

Lack of interaction between peripheral injection of CCK and obestatin in the regulation of gastric satiety signaling in rodents

Obestatin is a new peptide for which anorexigenic effects were recently reported in mice. We investigate whether peripheral injection of obestatin or co-injection with cholecystokinin (CCK) can modulate food intake, gastric motor function (intragastric pressure and emptying) and gastric vagal afferent activity in rodents. Obestatin (30, 100 and 300 microg/kg, i.p.) did not influence cumulative food intake for the 2h post-injection in rats or mice nor gastric emptying in rats. In rats, obestatin (300 microg/kg) did not modify CCK (1 microg/kg, i.p.)-induced significant decrease in food intake (36.6%) and gastric emptying (31.0%). Furthermore, while rats injected with CCK (0.3 microg/kg, i.v.) displayed gastric relaxation, no change in gastric intraluminal pressure was elicited by obestatin (300 microg/kg, i.v.) pre- or post-CCK administration. In in vitro rat gastric vagal afferent preparations, 20 units that had non-significant changes in basal activity after obestatin at 30 microg responded to CCK at 10 ng by a 182% increase. These data show that obestatin neither influences cumulative food intake, gastric motility or vagal afferent activity nor CCK-induced satiety signaling.     

Intratracheal injection of nickel chloride and copper-zinc superoxide dismutase activity in lung of rats.

Superoxide radical (O2-) is a free radical that may be involved in various toxic processes. Cu--Zn superoxide dismutase catalyses the dismutation of the superoxide free radical and protects cells from oxidative damage, and it has been used clinically. The concentration of Ni2+ and Cu--Zn superoxide dismutase activity were measured in lungs of rats at time intervals of 5, 12, 19, 26, 33, and 40 days following an intratracheal injection of 127 nmol of NiCl2. Nickel chloride increased nickel content and resulted in a significant increase of Cu--Zn superoxide dismutase activity in lungs. This elevation of Cu--Zn superoxide dismutase activity was highest on the 12th day (approximately threefold) and is at levels comparable to controls rats on day 40 onwards. Since Cu--Zn superoxide dismutase activity was increased in lung throughout our experimental period without corresponding increases of Cu2+ and Zn2+, we speculate that the elevation of Cu--Zn superoxide dismutase activity might be due to an increased half-life of the enzyme, induced by nickel.     

Aglomerular pathways in intrarenal microvasculature of aged rats

By means of silicone rubber injections, we confirmed the existence of several types of aglomerular arterial pathways within kidneys of aged rats. In superficial cortex some interlobular arteries divide to form aglomerular branches (Ludwig's arterioles) towards cortex corticis. In juxtamedullary cortex these pathways are relatively more numerous, they comprise: (a) Vasa Recta Vera, (b) glomeruli in which afferent and efferent arterioles form a continuous vessel and (c) glomeruli with two efferent vessels, one by-passing glomerular tuft. In addition, results obtained in the rat by the microsphere technique are in agreement with our morphological observations.     

Migration of monocytes after intracerebral injection

Recently, we monitored green fluorescent protein (GFP)-expressing monocytes after injection at the entorhinal cortex lesion (ECL) site in mice. We followed their migration out of the central nervous system (CNS) along olfactory nerve fibers penetrating the lamina cribrosa, within the nasal mucosa, and their subsequent appearance within the deep cervical lymph nodes (CLN), with numbers peaking at day 7. This is the same route activated T cells use for reaching the CLN, as we have shown before. Interestingly, GFP cells injected into the brain and subsequently found in the CLN exhibited ramified morphologies, which are typical of microglia and dendritic cells. To gain more insight into immunity and regeneration within the CNS we want to monitor injected monocytes using magnetic resonance imaging (MRI) after labeling with very small superparamagnetic iron oxide particles (VSOP). Due to their small size, nanoparticles have huge potential for magnetic labeling of different cell populations and their MRI tracking in vivo. So far we have verified that incubation with VSOP particles does not alter their migration pattern after ECL.     

Migration of monocytes after intracerebral injection

Recently, we monitored green fluorescent protein (GFP)-expressing monocytes after injection at the entorhinal cortex lesion (ECL) site in mice. We followed their migration out of the central nervous system (CNS) along olfactory nerve fibers penetrating the lamina cribrosa, within the nasal mucosa, and their subsequent appearance within the deep cervical lymph nodes (CLN), with numbers peaking at day 7. This is the same route activated T cells use for reaching the CLN, as we have shown before. Interestingly, GFP cells injected into the brain and subsequently found in the CLN exhibited ramified morphologies, which are typical of microglia and dendritic cells. To gain more insight into immunity and regeneration within the CNS we want to monitor injected monocytes using magnetic resonance imaging (MRI) after labeling with very small superparamagnetic iron oxide particles (VSOP). Due to their small size, nanoparticles have huge potential for magnetic labeling of different cell populations and their MRI tracking in vivo. So far we have verified that incubation with VSOP particles does not alter their migration pattern after ECL.     

Distribution of various nickel compounds in rat organs after oral administration

In this study, eight kinds of nickel (Ni) compounds were orally administered to Wistar male rats and the distribution of each compound was investigated 24 h after the administration. The Ni compounds used in this experiment were nickel metal [Ni−M], nickel oxide (green) [NiO(G)], nickel oxide (black) [NiO(B)], nickel subsulfide [Ni₃S₂], nickel sulfide [NiS], nickel sulfate [NiSO₄], nickel chloride [NiCl₂], and nickel nitrate [Ni(NO₃)₂]. The solubilities of the nickel compounds in saline solution were in the following order; [Ni(NO₃)₂>NiCl₂>NiSO₄]≫[NiS>Ni₃S₂]>[NiO(B)>Ni−M>NiO(G)]. The Ni level in the visceral organs was higher in the rats given soluble Ni compounds; Ni(NO₃)₂, NiCl₂, NiSO₄, than that in the rats receiving other compounds. In the rats to which soluble Ni compounds were administered, 80–90% of the recovered Ni amounts in the examined organs was detected in the kidneys. On the other hand, the Ni concentration in organs administered scarcely soluble Ni compounds; NiO(B), NiO(G), and Ni−M were very low. The estimated absorbed fraction of each Ni compounds was increased with the increase of the solubility. These results suggest that the kinetic behavior of Ni compounds administered orally is closely related with the solubility of Ni compounds, and that the solubility of Ni compounds is one of the important factors for determining the health effect of Ni compounds.