Aberrant histone modification in CD19⁺ B cells of patients with chronic lymphocytic leukemia

Figures & data

Table 1 Primer sequences

Name	Sequence 5′–3′
SIRT1 forward	TAC CGA GAT AAC CTT CTG
SIRT1 reverse	TCC AGT CAC TAG AGC TTG
HDAC1 forward	CAA GCT CCA CAT CAG TCC TTC C
HDAC1 reverse	TGC GGC AGC ATT CTA AGG TT
HDAC2 forward	AGT CAA GGA GGC GGC AAA A
HDAC2 reverse	TGC GGA TTC TAT GAG GCT TCA
HDAC7 forward	CTT CTC CAC AAG GAC AAG
HDAC7 reverse	CTC CAG GGT TCT GTA GG
PCAF forward	ATG AAT ATG CAA TTG GAT AC
PCAF reverse	CTC CTT CAT AAT CCT TGA TA
CREBBP forward	CTG CAC ACG ACA TGA CT
CREBBP reverse	GAA GTG GCA TTC TGT TG
P300 forward	CTG TAT GTG CTC CAG AAC
P300 reverse	GAC AAA AAG GCA GTT CC
EZH2 forward	ACATCCTGACTTCTGTGAG
EZH2 reverse	GGAGACCAAGAATACATTA
SUV39H1 forward	AAGAAGATCCGCGAACAGGAA
SUV39H1 reverse	GGAAGTGCTTGAGGATACGCAC
SUV39H2 forward	ATCCCACCTGGTACTCCCATCT
SUV39H2 reverse	GCAAAGCGAATACTGTGTGCC
β-actin forward	CCTGGCACCCAGCACAAT
β-actin reverse	GGGCCGGACTCGTCATAC

Table 2 Demographic and clinical characteristics of patients with CLL (n=58)

Characteristics	No of patients (%)
Sex
Male	34 (58.6)
Female	24 (41.4)
Age (years)
≤ 60	22 (37.9)
>60	36 (62.1)
Binet stage
A	15 (25.9)
B	20 (34.4)
C	23 (39.6)
Rai risk stage
Good	3 (5.2)
Intermediate	34 (58.6)
High	21 (36.2)
LDH
Normal	21 (36.2)
Elevated	37 (63.8)
β2-MG
Normal	23 (39.7)
Elevated	35 (60.3)
IgVH
Mutated	36 (62.1)
Unmutated	22 (37.9)
ZAP-70
Negative	48 (82.8)
Positive	10 (17.2)
CD38
Negative	47 (81.0)
Positive	11 (19.0)
FISH cytogenetics
No aberration or sole del(13q)	32 (55.2)
Del(17p) or del(11q) or trisomy 12	26 (44.8)

Abbreviations: CLL, chronic lymphocytic leukemia; FISH, fluorescence in situ hybridization; LDH, lactate dehydrogenase; β2-MG, β2 microglobulin.

Figure 1 Global histone H3/H4 acetylation and H3K9 methylation in subjects.

Notes: The global histone H3/H4 hypoacetylation in the CD19⁺ B cells of patients with CLL compared with controls (A and B) (P=0.028 and P=0.03, respectively). No significant difference in the global histone H3K4 methylation between the two groups (C). The global histone H3K9 methylation in patients with CLL was significantly higher than that of healthy controls (D) (P=0.02). The bars are presented as mean ± SD; OD units at 450 nm.
Abbreviations: CLL, chronic lymphocytic leukemia; OD, optical density; SD, standard deviation.

Figure 2 The SIRT1 mRNA expression in subjects.

Notes: The expression level of SIRT1 mRNA in patients with CLL was significantly higher than that of healthy controls (P=0.03) (A). The level of SIRT1 mRNA expression increased significantly with progression from Binet stage A to stage C (P=0.015) (B) and Rai good to high risk stage (P=0.007) (C). The bars indicate the relative mRNA levels of SIRT1 (mean ± SD).
Abbreviations: CLL, chronic lymphocytic leukemia; SD, standard deviation.

Figure 3 The EZH2 mRNA expression in subjects.

Notes: The expression level of EZH2 mRNA in patients with CLL was significantly higher than that of healthy controls (P=0.02) (A). The level of EZH2 mRNA expression increased significantly with progression from Binet stage A to stage C (P=0.01) (B) and Rai good to high risk stage (P=0.008) (C). The level of EZH2 mRNA expression increased significantly in the β2-MG+ subgroup compared with that in the β2-MG− subgroup (P=0.029) (D). The bars indicate the relative mRNA levels of SIRT1 (mean ± SD). β2-MG+, defined as β2-MG >3.5 mg/L; β2-MG−, defined as β2-MG ≤3.5 mg/L.
Abbreviations: CLL, chronic lymphocytic leukemia; β2-MG, β2 microglobulin; SD, standard deviation.

Figure 4 The mRNA expression of selected chromatin modifier genes in subjects.

Notes: The expression level of HDAC1 mRNA in patients with CLL was significantly higher than that of healthy controls (P=0.02) (A). The level of HDAC2 mRNA expression was reduced significantly in patients with CLL (P=0.002) (B), HDAC7 mRNA expression level was increased in patients with CLL (P=0.008) (C). The level of P300 mRNA expression was reduced significantly in patients with CLL (P=0.001) (D). The bars indicate the relative mRNA levels of SIRT1 (mean ± SD).
Abbreviations: CLL, chronic lymphocytic leukemia; SD, standard deviation.

Figure 5 The mRNA expression of selected chromatin modifier genes in subjects.

Note: There was no significant difference in the expression of PCAF (A), CREBBP (B), SUV39H1 (C), and SUV39H2 (D) mRNA between patients with CLL and controls, and between any subgroups.
Abbreviation: CLL, chronic lymphocytic leukemia.