**8. Conclusion**

114 Blood Cell – An Overview of Studies in Hematology

evaluation of physiological state.

Stresses

Number of genes

subpopulations.

**6. Gene expression profiles change with other stresses** 

Furthermore, a possibility was shown that whole blood RNA analysis is applicable to

The degree of stress can be comparable according to the numbers of up-regulated and

Sodium azide was given orally to the miniature pigs over 20 weeks. There were no significant changes of hematological and biochemical properties for administrated dose of 300µg/kg, one hundredth of LD50. On the other hand, gene expression profiles were obviously changed. Anesthesia group showed a slight degree, but the one week fasting group showed a significant difference. This can be clearly noticed when the contents of stress is classified by the function of up-regulated and down-regulated genes. Consequently,

sodium azide 300µg/kg ; LD50 1/100 893 339 554 blood removal(150ml)after 6 hours 1747 227 1520

**7. Effects of white blood cells on whole blood gene expression profiles** 

Whole blood contains a variety of cell types as red blood cells, granulocytes, lymphocytes, and platelets. Most of the nucleated cells in blood are white blood cells such as neutrophils, T-cells, B-cells, and monocytes. The number of white blood cells in humans is known to decrease steadily from infancy to adulthood, and its composition (i.e. lymphocytes, granulocytes) also changes with age [30]. In study of the gene expression profiles change related to aging, hematological data of the fetal stage was unavailable because the amount of collected blood was insufficient for the analysis. From 12 to 30 weeks of age, ANOVA analysis indicated no significant differences in the fractions of lymphocytes, neutrophils, eosinophils, basophils, and monocytes. In addition, these compositions were almost equal to those in human adults. The above result suggests that the gene expression profile change of age-related whole blood RNA is not due to the composition of white blood cell

The intraclass correlation between Staphylococcus enterotoxin B-stimulated and unstimulated blood from healthy subjects was significantly higher in leukocyte-derived samples the in whole blood, suggesting that the method of RNA isolation from whole blood

Fasting a week 3136 1840 1296 anesthesia after 6 hours 160 87 73 non treatment (blood removal 20ml) 73 14 59

P<0.05, Fold change>2 total up regulation down

regulation

down-regulated genes, even if the stress is different in quality from the others.

grade of the stress can be estimated according to the expression state of genes.

**Table 4.** Summery of gene expression condition of several types of stress

Whole blood RNA is easy to handle compared to isolated white blood cell RNA and can be used for health and disease monitoring and animal control. In addition, whole blood is a heterogeneous mixture of subpopulation cells. Once a great change occurs in composition and expressing condition of subpopulations, their associated change will be reflected on whole blood RNA.

Whole blood microarray analyses were conducted to evaluate variations of correlation among individuals and ages using specific pathogen-free (SPF) Clawn miniature pigs. The characteristics of age-related gene expression by taking into account of change in the number of expressed genes by age and similarities of gene expression intensity between individuals were identified. As a result, the number of expressed genes was less in fetal stage and infancy period but increased with age, reaching a steady state of gene expression after 20 weeks of age. Variation in gene expression intensity within the same age was great in fetal stage and infancy period, but converged with age. The variation between 20 and 30

weeks of age was comparable to that among 30 weeks individuals. These results indicate that uniformity of laboratory animals is expected for miniature pigs after 20 weeks of age.

Whole Blood RNA Analysis, Aging and Disease 117

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In dietary-induced hyperlipidemia study, feeding treatments commenced when the pigs were 12 weeks old, RNA analysis was conducted on whole blood sampled after 10, 19, and 27 weeks of the feeding period. Variation in whole blood gene expression intensity among individuals within the HFCD group was in the same range as that of the controls at any period, indicating uniformity of dietary-induced hyperlipidemia expression profiles in miniature pigs. Dietaryinduced transitions of gene expression profiles for genes bearing GO terms were examined. Major changes included an induction of proteins involved in catabolic processes and protein metabolism after a 19-week dietary period, and a reduced expression of proteins involved in steroid metabolism and lipid biosynthesis after a 27-week dietary period.

In several kinds of stress study, the degree (extent) of stress can be comparable according to the gene number of up-regulate, or down-regulate, even if the stress is different in kind from the others.

A possibility was shown that whole blood RNA analysis is applicable to evaluation of physiological state. By considering variation in gene expression profiles of miniature pigs, whole blood RNA analyses can be used in practical applications. The blood RNA diagnostics under development may eventually be useful for monitoring human health.
