Effect Of SN56 Cells' Livability To Low Concentration Of Aβ (1-42)monomer/Oligomers And CO Donor

Alzheimer's disease (AD) is a kind of neurodegenerative disease, and associated with cognitive dysfunction and memory loss as the main clinical symptoms. It has been an important disease which was threatening the health of the old now. And its morbidity rate increases every year. 1906, the typical clinical symptoms and pathological features of AD were firstly demonstrated thoroughly by Alois Alzheimer. But until now, we still know little about its exact pathological mechanism. AD is characterized by its anomalous self-assembled and deposited ofβ-amyloid peptides(Aβ) in the neuritic plaques and neurofibrillar tangles. Aβ(1-42) is the major amyloid component in the neuritic plaques of the affected brain, indicating a key role of Aβ(1-42) in amyloidogenesis and the associated neurobehavioral impairments in AD. The form in which Aβ(1-42) is existed and the concentration it needs to facilitat AD invasion remain an controversial question and are the hot spots between researchers in recent years, at home and abroad. Endogenous carbon monoxide (CO) is an important chemical neurotransmitter in the body. It has the role of anti-apoptosis in certain cell types, but its functions on nerve cells remain unknown. Recently, the transitional metal carbonyls have been identified as potential CO-releasing molecules (CORMs) on account of its ability to facilitate the pharmaceutical use of CO by delivering it to tissues and organs. It contains a variety of heavy metals such as cobalt, iron, nickel and ruthenium. CO, releases from these metals can be modulated by photoactivation, temperature, pH and the nature of ambient environment. The tricarbonyldichlor oruthenium (â…¡) dimer has been confirmed as one of the novel group of CORMs. It is usually termed as CORM-2. Many researches have suggested that it could play a pivotal role of anti-apoptosis in some cells. Consequently, it has been treated as a kind of donor for CO, and been widely used in experimental researches. Objective :1 To observe the susceptibility of SN56 cells to Aβ(1-42) monomer/oligomers in low concentration through examining the effect of SN56 cells'liviability to low concentration of Aβ(1-42) monomer/oligomers.2 To observe weather CORM-2 could play a protective role on SN56 through examining the liviability variation of SN56 cells when cultured them in the environment of Aβ(1-42) monomer/oligomers and different concentrations of CORM-2.Methods:1 Cell cultureSN56 cells were incubated in a 37℃, 95% air, 5% CO2 environment. A EDTA-Ca²⁺-Trypsin in the concentration of 0.05% was used to the cell's digestion and passage. Plated cells onto the 96-well plate with uniform concentration. SN56 Cells were grown for 3 days before they proliferated and adhered completely. Differentiation was achieved by culture with 1mM cyclic adenosine monophosphate (cAMP) and 1μM retinoic acid (RA) for 3 days. Differentiated SN56 cells were harvested finally.2 GroupingThe cells were divided into four groups, and named as: control group, Aβ(1-42)group, Aβ(1-42)+CORM-2 50μM group, Aβ(1-42)+CORM-2 100μM group. The first three columns of the 96-well plate were the control group which didn't contain any effect factors, and the others were added Aβ(1-42)10nM, Aβ(1-42) 100nM, Aβ(1-42)1μM, Aβ(1-42)10nM+CORM-2 50μM, Aβ(1-42) 100nM+CORM-2 50μM, Aβ(1-42)1μM+CORM-2 50μM, Aβ(1-42) 10nM+CORM-2 100μM, Aβ(1-42)100nM+CORM-2 100μM, Aβ(1-42)1μM+ CORM-2 100μM separately to each column in sequence. Assayed the liviability of each group after 3 days culture.3 MTT cell survival assayAdded 11μl of MTT solution to each well of 96-well plate. Incubated at 37℃in 5% CO2 for 3 hours. Added 111μl mixture of isopropyl alcohol and hydrogen chloride for each well. pumped repeatedly to mix them and laied aside for 2⁵mins. Assayed the absorbancy of each well at 570nm wavelength.4 Statistical MethodsThe results were recorded as x±s. Data processing was done by SPSS13.0 statistical software. Multiple independent samples were compared by Kruskal-Wallis test, two-related samples were compared by Wilcoxon test. P<0.05 had statistically significant.Results :The liviability of Aβ(1-42) group are (79.73±0.94)%, (67.99±0.79)%, (60.42±0.62)%, respectively,decreasing with the concentration of Aβ(1-42) increasing. When cocultured with CORM-2 50μM, the liviabilities are (75.15±0.096)%, (63.20±0.17)%, (55.33±0.19)%, lower than Aβ(1-42) group respectively. When cocultured with CORM-2 100μM, the liviabilities are (73.20±0.27)%, (64.34±0.11)%, (54.17±0.12)%, also lower than Aβ(1-42) group, but has discrepancy in the ability of decreasing the cell livability when compared with Aβ(1-42) + CORM-2 50μM group.Conclusion:Low concentration of Aβ(1-42) monomer/oligomers could reduce the liviability of SN56 cells, and higher concentration has more significant effect, indicating that its effect on SN56 cells is closely related with the concentration; different concentrations of CORM-2 could reduce SN56 cells'livability, and there is a discrepancy in the intensity. Therefore, we propose that a certain concentration of CORM-2 couldn't protect SN56 cells form toxic damage. On the contrary, it aggravates the cells to die. However, the relationship between the intensity and dose need to be clarified further.