893
Annals of the Rheumatic Diseases 1990; 49: 893-895
Enhanced expression of the heat shock protein gene in peripheral blood mononuclear cells of patients with active systemic lupus erythematosus Y Deguchi, S Kishimoto
Abstract The spontaneous increase in the transcription of the heat shock protein (hsp 70) gene in peripheral blood mononuclear cells of patients with active systemic lupus erythematosus (SLE) is shown by nuclear run on transcription assay. The transcription of hsp 70 gene in the peripheral blood mononuclear cells of five patients with active SLE was more than 10 times greater than that in five normal healthy subjects or three patients with bronchial asthma as controls. This suggests that heat shock proteins may be produced during an active immune response in patients with active SLE and play a part in a change related to lupus of the essential intracellular functions of peripheral blood mononuclear cells.
Third Department of Internal Medicine, Osaka University School of Medicine, Fukushima-ku, Osaka 553, Japan Y Deguchi S Kishimoto
Correspondence to: Dr Yasuhiro Deguchi, Laboratory of Immunoregulation, National Institute of Allergy and Infectious Diseases, NIH, 10687 Weymouth St No 103, Bethesda, Maryland 20814, USA. Accepted for publication 5 December 1989
Systemic autoimmune disorders, such as systemic lupus erythematosus (SLE), are characterised by immunological dysfunction, affecting skin, lung, heart, and muscle.' Investigators have directed attention towards immunological abnormalities,l and there are few data about the pathophysiology of intracellular molecules of peripheral blood mononuclear cells in autoimmune diseases. Changes of gene expression in response to heat shock stress (raised temperature) have been described in animal cells.2 3 The general effect of heat shock stress is the suppression of protein synthesis, normally produced at a normal temperature, and enhancement and further synthesis of new proteins such as heat shock proteins (hsps). Schlesinger et al showed that various chemical, mechanical and environmental stresses could induce hsps.4 It has been suggested that these proteins might be important for cell survival under environmental or physiological stresses.5 6 The 70 kD hsp (hsp 70) family includes the inducible and cognate 68 to 72 kD hsps; hsp 70 is the most conserved and well characterised. It has been suggested recently that hsps are also developmentally regulated
and may have an important and essential role in proliferation and differentiation.7 The half
cell
life of RNA transcipt for hsp 70 is short.7
In this study we examined the transcriptional level of the hsp 70 gene in peripheral blood mononuclear cells of patients with active SLE by nuclear run on transcription assay.
Patients, materials, and methods PATIENTS AND CONTROLS
We examined five patients (four female, one male) with active SLE. All were receiving prednisolone or azathioprine, or both, and all met the American Rheumatism Association criteria for SLE.8 The table gives clinical and laboratory data for these patients. We also examined three patients with bronchial asthma who had been receiving 20 mg or more of prednisolone daily for the treatment of asthmatic attacks, and five healthy volunteers who were laboratory or hospital personnel and had no history of use of any drugs known to affect immune functions.
PREPARATION OF PERIPHERAL BLOOD MONONUCLEAR CELLS Heparinised peripheral blood was diluted two-
fold with RPMI 1640 medium (Bioproducts Inc, USA), layered on Ficoll-Paque, and centrifuged at 1800 rpm for 20 minutes. Peripheral blood mononuclear cells were collected from the interface and washed twice with RPMI 1640 medium. The cell population was over 97% viable (trypan blue exclusion). NUCLEAR RUN ON TRANSCRIPTION ASSAY
To obtain nuclei cells were lysed in a solution containing 10 mM TRIS (pH 7-5), 2 mM MgCI2, 3 mM CaCl2, 5 mM dithiothreitol, and 0-02% nonidet P-40, with subsequent centrifugation through 2 M sucrose solution. Thirty million nuclei were suspended in 100 tl of
Clinical and laboratory data for the autoimmune patients ixn this study Patient No
Sex
I 2 3 4 5
F M F F F
Clinical activity*
Clinical
manifestationst
Autoantibodies
ANFt Anti-dsDNA5 (Ulnd) 210 Sk, R, N, H, F 122 Se, R, A, F 210 64 Sk, O, A, H, F 28 64 Sk, R, N, H, F 210 122 Sk, O, A, H, F, P 210 64 *Clinical activity was determined according to the UCH/Middlesex criteria.9 fA=arthritis; F=high fever; H=haematological disorder; N=neurological disorder; O=oral ulcer; P=pulmonary disorder; R=renal disorder; Se=serositis; Sk=skin disorder. tTitre of antinuclear factor (ANF) is less than 2 in normal subjects. Very active Active Active Very active Very active
SAnti-dsDNA (anti-double-stranded DNA) is less than 10 U/ml in normal subjects.
894
^
Deguchi, Kishimoto
50% glycerol solution with 50 mM TRIS (pH 7 5), 5 mM MgCl2, and 0 mM EDTA. The suspension of nuclei was immediately mixed with an equal volume of buffer containing 0-2 M KC1, 5 mM MgCl2, 5 mM dithiothreitol, 1 mM ATP, CTP, GTP, and 200 units of RNasin (ribonuclease inhibitor, 500 units; Amersham International plc, England). The preparation was then incubated at 28°C for 20 minutes after addition of 1-85 MBq of 32P radiolabelled UTP (111 GBq/ml; Amersham Inc). Sodium dodecyl sulphate (SDS) and EDTA solution were added to a final concentration of 1% and 5 mmol/I respectively, followed by treatment with proteinase K (1 mg/ml) at 42°C for 30 minutes. RNA was extracted with phenol and chloroform from the preparation and precipitated with ethanol. The pellet was resuspended in 3 ml of hybridisation buffer, which contained 50% formamide, 0-75 M NaCl, 0-5% SDS, 2 mM EDTA, 50 mM HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulphonic acid) (pH 7 0), one tenth dilution of Denhardt's solution, and denatured salmon sperm DNA (500 ,ug/ml).'° Finally, the preparation was applied to the nitrocellulose filter onto which the human hsp 70 cDNA probe (1 2 kb, Bam HI fragment) or ,B actin probe (Wako Pure Chemical Industries, Japan)" had been dotted. After 24 hours' incubation the filter was washed three times in
SLE
-
O0IxSSC (0 15 M NaCl+0015 M sodium citrate) and 0- 1% of SDS at 60°C and appropriate solutions, dried, and exposed to x ray film with intensifying screen at -70°C. In some experiments the hybridised dots were excised from the filter and directly counted by betacounter.12
o_
Normal controls
0
20 10 HSP70 transcription (relative counts /10 5 cells)
Figure 2 Measurement ofhsp 70 gene trasnscription in peripheral blood mononuclear cells ofpatients with active systemic lupus erythematosus (S), patients with bronchial asthma (B), and normal healthy subjects (N). The relative amount (relative counts/llO cells) ofhsp 70 gene transcription was determined with a betacounter for the hybridised dots in the nuclear run on transcription assay. The mean values and standard deviations are shown.
healthy subjects as controls. We used a nuclear run on transcription technique with human hsp 70 cDNA and a ,B actin probe. We first found a spontaneous increase of the transcriptional level of the hsp 70 gene in peripheral blood mononuclear cells of patients with active SLE (fig 1). The use of the I actin probe provided good control as actin synthesis is unaffected by heat stress. We found no significant change in the transcriptional level of the human actin gene in them. We further measured the hybridisation signal by betacounter; fig 2 summarises the transcriptional level of the hsp 70 gene in peripheral blood mononuclear cells of patients with active SLE and control subjects. The transcriptional level of the hsp 70 gene in peripheral blood mononuclear cells of patients with active SLE was found to be more than 10 times greater than that of patients with bronchial asthma or that of normal healthy subjects.
STATISTICAL ANALYSIS
Statistical analysis of the data was by Student's t Discussion Various chemical, mechanical, and environmental stresses, including heat shock response, can induce the transcription of the hsp gene.' Results We compared the transcriptional level of the Heat shock 70 kD and 85 kD (hsp 70 and hsp hsp 70 gene in peripheral blood mononuclear 85) are major components of human cells.7 In cells of patients with active SLE with that in this study we have shown the spontaneous patients with bronchial asthma and normal activation of hsp 70 gene transcription in peripheral blood mononuclear cells of patients with active SLE. In eukaryotes transcription of the heat shock protein gene has been most 2 3 4 intensively investigated in drosophila. 14 The 1 activation of hsp gene transcription is mediated by specific sequence elements in the heat shock promoters. Deletion analyses showed that 20 base pairs located about 20 nucleotides upA .e stream from the 5' portion to the TATA box were essential for induction of the hsp 70 gene in drosophila.'5 The sequences were recognised by multimeric DNA binding proteins.'6 It is test.
B
; .~-
- s
~not clear how the spontaneous activation of transcription of the hsp 70 gene in peripheral
- v
Figure I Representative result of nuclear run on transcription analysisfor hsp 70 (A) (48 hour exposure autoradiogram) and ( actin gene (B) expression (12 hour exposure autoradiogram). Lanes I and 2 denote peripheral blood mononuclear cells of subjects with active systemic lupus erythematosus (cases 2 and 5), lane 3 denotes those of a patient with bronchial asthma, and lane 4 those of a normal healthy subject.
blood mononuclear cells of patients with active SLE is regulated by the similar DNA binding
proteins.
As hsps are not well known to
participate in peripheral blood mononuclear cell functions it is possible that they play an essential part themntracellular mechanisms of peripheral blood mononuclear cell functions. Hsps are also developmentally regulated and
Increased transcriptin ofheat shock protein gene in SLE
play a part in cell differentiation and proliferation.'7 Interleukin-2 and mitogens increase hsp 70 mRNA in lymphocytes.'8 It has been shown that hsps indeed participate in inflammation-for example, in severe osteoarthritis. 9 We also reported that at protein level the amount of some hsps increased in peripheral blood mononuclear cells from patients with SLE.20 As a result of the present study enhanced transcription of the hsp gene is clearly a possible mechanism for the increased amount of hsp products at protein level in peripheral blood mononuclear cells, of patients with active SLE. An understanding of the intracellular events in the peripheral blood mononuclear cells of patients with active SLE involved in hsp gene activation, and of the role of hsp products themselves, may provide some new approaches to determining the pathophysiology of chronic inflammation processes such as SLE. The essential significance of the spontaneous activation of the hsp gene in the peripheral blood mononuclear cells of patients with active SLE is in progress. We thank Dr Ariga (Hokkaido University) for the generous gift of the human hsp 70 cDNA probe. This study was supported in part by grants from the Ministry of Culture and Education and Science, Japan. Jiminez S A. Cellular immune dysfunction and the pathogenesis of scleroderma. Semin Arthritis Rheun 1983; 13: 104-20. 2 Hichey E D, Weber L A. Modulation of heat shock polypeptide synthesis in Hela cells during hyperthermia and recovery. Biochemistry 1982; 21: 1513-21. 3 Morange M, Din A, Bensude 0, Babinet C. Altered 1
895 expression ot heat shock proteins in embryonal carcinoma and mouse early embryonic cells. Mol Cell Biol 1984; 4: 730-5. 4 Schlesinger M J, Aliperti G, Kelly P M. The response of cells to heat shock. Trends in Biochemical Science 1982; 7: 222-5. 5 Currie R W, White F P. Trauma-induced protein in rat tissues. Science 1981; 214: 72-3. 6 Lindquist S. The heat shock response. Annu Rev Biochem 1986; 55: 1151-91. 7 Lindquist S, Craig E A. The heat shock response. Annu Rev Genet 1988; 22: 631-77. 8 Tan E M, Cohen A S, Fries J F, et al. The 1982 revised criteria for the classification of systemic lupus erythematosus. Arthritis Rheum 1982; 25: 1271-7. 9 Morrow W J W, Isenberg D A, Parry H F, Snaith M L. Creactive protein in sera from patients with systemic lupus erythematosus. J Rheunatol 1981; 8: 599-604. 10 Marzluff W F, Huang R C C. Transcription and translation, a practical approach. Oxford: IRL Press, 1984: 89-130. 11 Deguchi Y, Negoro S, Kishimoto S. Age-related changes of heat shock protein gene transcription in human peripheral blood mononuclear cells. Biochem Biophys Res Commun 1988; 157: 580-4. 12 Reed J C, Tsujimoto Y, Alpers J D, Croce C M, Nowell P C. Regulation of bcl-2 protooncogene expression during normal human lymphocyte proliferation. Science 1987; 236: 1295-9. 13 Anathan J, Goldberg A L, Voellmy R. Abnormal proteins serve as eukaryotic stress signals and trigger the activation of heat shock genes. Science 1988; 232: 5224. 14 Lindquist S. Varying pattern of protein synthesis in drosophila during heat shock. Dev Biol 1980; 77: 463-79. 15 Pelham H R B. A regulatory upstream promoter element in the drosophila hsp 70 heat shock gene. Cell 1982; 30: 517-28. 16 Parker C S, Topol J. A drosophila RNA polymerase II transcription factor binds to the regulatory site of an hsp 70 gene. Cell 1984; 37: 273-83. 17 Kingston R E, Baldwin A S, Sharp P A. Regulation of heat shock protein 70 gene expression by c-myc. Nature 1984; 315: 280-2. 18 Granelli-Piperno A, Andrus L, Steinman R M. Lymphokine and non lymphokine mRNA levels in stimulated human T cells. J Exp Med 1986; 163: 922-37. 19 Kubo T, Towele C A, Markin H J, Treadwell B V. Stressinduced proteins in chondrocytes from patients with osteoarthritis. Ardhiis Rhewn 1985; 28: 1140-5. 20 Deguchi Y, Negoro S, Kishimoto S. Heat shock protein synthesis in peripheral blood mononuclear cells from patients with systemic lupus erythematosus. Biochem Biophys Res Commun 1987; 148: 1063-8.
Annals of the Rheumatic Diseases 1990; 49: 893-895
Enhanced expression of the heat shock protein gene in peripheral blood mononuclear cells of patients with active systemic lupus erythematosus Y Deguchi, S Kishimoto
Abstract The spontaneous increase in the transcription of the heat shock protein (hsp 70) gene in peripheral blood mononuclear cells of patients with active systemic lupus erythematosus (SLE) is shown by nuclear run on transcription assay. The transcription of hsp 70 gene in the peripheral blood mononuclear cells of five patients with active SLE was more than 10 times greater than that in five normal healthy subjects or three patients with bronchial asthma as controls. This suggests that heat shock proteins may be produced during an active immune response in patients with active SLE and play a part in a change related to lupus of the essential intracellular functions of peripheral blood mononuclear cells.
Third Department of Internal Medicine, Osaka University School of Medicine, Fukushima-ku, Osaka 553, Japan Y Deguchi S Kishimoto
Correspondence to: Dr Yasuhiro Deguchi, Laboratory of Immunoregulation, National Institute of Allergy and Infectious Diseases, NIH, 10687 Weymouth St No 103, Bethesda, Maryland 20814, USA. Accepted for publication 5 December 1989
Systemic autoimmune disorders, such as systemic lupus erythematosus (SLE), are characterised by immunological dysfunction, affecting skin, lung, heart, and muscle.' Investigators have directed attention towards immunological abnormalities,l and there are few data about the pathophysiology of intracellular molecules of peripheral blood mononuclear cells in autoimmune diseases. Changes of gene expression in response to heat shock stress (raised temperature) have been described in animal cells.2 3 The general effect of heat shock stress is the suppression of protein synthesis, normally produced at a normal temperature, and enhancement and further synthesis of new proteins such as heat shock proteins (hsps). Schlesinger et al showed that various chemical, mechanical and environmental stresses could induce hsps.4 It has been suggested that these proteins might be important for cell survival under environmental or physiological stresses.5 6 The 70 kD hsp (hsp 70) family includes the inducible and cognate 68 to 72 kD hsps; hsp 70 is the most conserved and well characterised. It has been suggested recently that hsps are also developmentally regulated
and may have an important and essential role in proliferation and differentiation.7 The half
cell
life of RNA transcipt for hsp 70 is short.7
In this study we examined the transcriptional level of the hsp 70 gene in peripheral blood mononuclear cells of patients with active SLE by nuclear run on transcription assay.
Patients, materials, and methods PATIENTS AND CONTROLS
We examined five patients (four female, one male) with active SLE. All were receiving prednisolone or azathioprine, or both, and all met the American Rheumatism Association criteria for SLE.8 The table gives clinical and laboratory data for these patients. We also examined three patients with bronchial asthma who had been receiving 20 mg or more of prednisolone daily for the treatment of asthmatic attacks, and five healthy volunteers who were laboratory or hospital personnel and had no history of use of any drugs known to affect immune functions.
PREPARATION OF PERIPHERAL BLOOD MONONUCLEAR CELLS Heparinised peripheral blood was diluted two-
fold with RPMI 1640 medium (Bioproducts Inc, USA), layered on Ficoll-Paque, and centrifuged at 1800 rpm for 20 minutes. Peripheral blood mononuclear cells were collected from the interface and washed twice with RPMI 1640 medium. The cell population was over 97% viable (trypan blue exclusion). NUCLEAR RUN ON TRANSCRIPTION ASSAY
To obtain nuclei cells were lysed in a solution containing 10 mM TRIS (pH 7-5), 2 mM MgCI2, 3 mM CaCl2, 5 mM dithiothreitol, and 0-02% nonidet P-40, with subsequent centrifugation through 2 M sucrose solution. Thirty million nuclei were suspended in 100 tl of
Clinical and laboratory data for the autoimmune patients ixn this study Patient No
Sex
I 2 3 4 5
F M F F F
Clinical activity*
Clinical
manifestationst
Autoantibodies
ANFt Anti-dsDNA5 (Ulnd) 210 Sk, R, N, H, F 122 Se, R, A, F 210 64 Sk, O, A, H, F 28 64 Sk, R, N, H, F 210 122 Sk, O, A, H, F, P 210 64 *Clinical activity was determined according to the UCH/Middlesex criteria.9 fA=arthritis; F=high fever; H=haematological disorder; N=neurological disorder; O=oral ulcer; P=pulmonary disorder; R=renal disorder; Se=serositis; Sk=skin disorder. tTitre of antinuclear factor (ANF) is less than 2 in normal subjects. Very active Active Active Very active Very active
SAnti-dsDNA (anti-double-stranded DNA) is less than 10 U/ml in normal subjects.
894
^
Deguchi, Kishimoto
50% glycerol solution with 50 mM TRIS (pH 7 5), 5 mM MgCl2, and 0 mM EDTA. The suspension of nuclei was immediately mixed with an equal volume of buffer containing 0-2 M KC1, 5 mM MgCl2, 5 mM dithiothreitol, 1 mM ATP, CTP, GTP, and 200 units of RNasin (ribonuclease inhibitor, 500 units; Amersham International plc, England). The preparation was then incubated at 28°C for 20 minutes after addition of 1-85 MBq of 32P radiolabelled UTP (111 GBq/ml; Amersham Inc). Sodium dodecyl sulphate (SDS) and EDTA solution were added to a final concentration of 1% and 5 mmol/I respectively, followed by treatment with proteinase K (1 mg/ml) at 42°C for 30 minutes. RNA was extracted with phenol and chloroform from the preparation and precipitated with ethanol. The pellet was resuspended in 3 ml of hybridisation buffer, which contained 50% formamide, 0-75 M NaCl, 0-5% SDS, 2 mM EDTA, 50 mM HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulphonic acid) (pH 7 0), one tenth dilution of Denhardt's solution, and denatured salmon sperm DNA (500 ,ug/ml).'° Finally, the preparation was applied to the nitrocellulose filter onto which the human hsp 70 cDNA probe (1 2 kb, Bam HI fragment) or ,B actin probe (Wako Pure Chemical Industries, Japan)" had been dotted. After 24 hours' incubation the filter was washed three times in
SLE
-
O0IxSSC (0 15 M NaCl+0015 M sodium citrate) and 0- 1% of SDS at 60°C and appropriate solutions, dried, and exposed to x ray film with intensifying screen at -70°C. In some experiments the hybridised dots were excised from the filter and directly counted by betacounter.12
o_
Normal controls
0
20 10 HSP70 transcription (relative counts /10 5 cells)
Figure 2 Measurement ofhsp 70 gene trasnscription in peripheral blood mononuclear cells ofpatients with active systemic lupus erythematosus (S), patients with bronchial asthma (B), and normal healthy subjects (N). The relative amount (relative counts/llO cells) ofhsp 70 gene transcription was determined with a betacounter for the hybridised dots in the nuclear run on transcription assay. The mean values and standard deviations are shown.
healthy subjects as controls. We used a nuclear run on transcription technique with human hsp 70 cDNA and a ,B actin probe. We first found a spontaneous increase of the transcriptional level of the hsp 70 gene in peripheral blood mononuclear cells of patients with active SLE (fig 1). The use of the I actin probe provided good control as actin synthesis is unaffected by heat stress. We found no significant change in the transcriptional level of the human actin gene in them. We further measured the hybridisation signal by betacounter; fig 2 summarises the transcriptional level of the hsp 70 gene in peripheral blood mononuclear cells of patients with active SLE and control subjects. The transcriptional level of the hsp 70 gene in peripheral blood mononuclear cells of patients with active SLE was found to be more than 10 times greater than that of patients with bronchial asthma or that of normal healthy subjects.
STATISTICAL ANALYSIS
Statistical analysis of the data was by Student's t Discussion Various chemical, mechanical, and environmental stresses, including heat shock response, can induce the transcription of the hsp gene.' Results We compared the transcriptional level of the Heat shock 70 kD and 85 kD (hsp 70 and hsp hsp 70 gene in peripheral blood mononuclear 85) are major components of human cells.7 In cells of patients with active SLE with that in this study we have shown the spontaneous patients with bronchial asthma and normal activation of hsp 70 gene transcription in peripheral blood mononuclear cells of patients with active SLE. In eukaryotes transcription of the heat shock protein gene has been most 2 3 4 intensively investigated in drosophila. 14 The 1 activation of hsp gene transcription is mediated by specific sequence elements in the heat shock promoters. Deletion analyses showed that 20 base pairs located about 20 nucleotides upA .e stream from the 5' portion to the TATA box were essential for induction of the hsp 70 gene in drosophila.'5 The sequences were recognised by multimeric DNA binding proteins.'6 It is test.
B
; .~-
- s
~not clear how the spontaneous activation of transcription of the hsp 70 gene in peripheral
- v
Figure I Representative result of nuclear run on transcription analysisfor hsp 70 (A) (48 hour exposure autoradiogram) and ( actin gene (B) expression (12 hour exposure autoradiogram). Lanes I and 2 denote peripheral blood mononuclear cells of subjects with active systemic lupus erythematosus (cases 2 and 5), lane 3 denotes those of a patient with bronchial asthma, and lane 4 those of a normal healthy subject.
blood mononuclear cells of patients with active SLE is regulated by the similar DNA binding
proteins.
As hsps are not well known to
participate in peripheral blood mononuclear cell functions it is possible that they play an essential part themntracellular mechanisms of peripheral blood mononuclear cell functions. Hsps are also developmentally regulated and
Increased transcriptin ofheat shock protein gene in SLE
play a part in cell differentiation and proliferation.'7 Interleukin-2 and mitogens increase hsp 70 mRNA in lymphocytes.'8 It has been shown that hsps indeed participate in inflammation-for example, in severe osteoarthritis. 9 We also reported that at protein level the amount of some hsps increased in peripheral blood mononuclear cells from patients with SLE.20 As a result of the present study enhanced transcription of the hsp gene is clearly a possible mechanism for the increased amount of hsp products at protein level in peripheral blood mononuclear cells, of patients with active SLE. An understanding of the intracellular events in the peripheral blood mononuclear cells of patients with active SLE involved in hsp gene activation, and of the role of hsp products themselves, may provide some new approaches to determining the pathophysiology of chronic inflammation processes such as SLE. The essential significance of the spontaneous activation of the hsp gene in the peripheral blood mononuclear cells of patients with active SLE is in progress. We thank Dr Ariga (Hokkaido University) for the generous gift of the human hsp 70 cDNA probe. This study was supported in part by grants from the Ministry of Culture and Education and Science, Japan. Jiminez S A. Cellular immune dysfunction and the pathogenesis of scleroderma. Semin Arthritis Rheun 1983; 13: 104-20. 2 Hichey E D, Weber L A. Modulation of heat shock polypeptide synthesis in Hela cells during hyperthermia and recovery. Biochemistry 1982; 21: 1513-21. 3 Morange M, Din A, Bensude 0, Babinet C. Altered 1
895 expression ot heat shock proteins in embryonal carcinoma and mouse early embryonic cells. Mol Cell Biol 1984; 4: 730-5. 4 Schlesinger M J, Aliperti G, Kelly P M. The response of cells to heat shock. Trends in Biochemical Science 1982; 7: 222-5. 5 Currie R W, White F P. Trauma-induced protein in rat tissues. Science 1981; 214: 72-3. 6 Lindquist S. The heat shock response. Annu Rev Biochem 1986; 55: 1151-91. 7 Lindquist S, Craig E A. The heat shock response. Annu Rev Genet 1988; 22: 631-77. 8 Tan E M, Cohen A S, Fries J F, et al. The 1982 revised criteria for the classification of systemic lupus erythematosus. Arthritis Rheum 1982; 25: 1271-7. 9 Morrow W J W, Isenberg D A, Parry H F, Snaith M L. Creactive protein in sera from patients with systemic lupus erythematosus. J Rheunatol 1981; 8: 599-604. 10 Marzluff W F, Huang R C C. Transcription and translation, a practical approach. Oxford: IRL Press, 1984: 89-130. 11 Deguchi Y, Negoro S, Kishimoto S. Age-related changes of heat shock protein gene transcription in human peripheral blood mononuclear cells. Biochem Biophys Res Commun 1988; 157: 580-4. 12 Reed J C, Tsujimoto Y, Alpers J D, Croce C M, Nowell P C. Regulation of bcl-2 protooncogene expression during normal human lymphocyte proliferation. Science 1987; 236: 1295-9. 13 Anathan J, Goldberg A L, Voellmy R. Abnormal proteins serve as eukaryotic stress signals and trigger the activation of heat shock genes. Science 1988; 232: 5224. 14 Lindquist S. Varying pattern of protein synthesis in drosophila during heat shock. Dev Biol 1980; 77: 463-79. 15 Pelham H R B. A regulatory upstream promoter element in the drosophila hsp 70 heat shock gene. Cell 1982; 30: 517-28. 16 Parker C S, Topol J. A drosophila RNA polymerase II transcription factor binds to the regulatory site of an hsp 70 gene. Cell 1984; 37: 273-83. 17 Kingston R E, Baldwin A S, Sharp P A. Regulation of heat shock protein 70 gene expression by c-myc. Nature 1984; 315: 280-2. 18 Granelli-Piperno A, Andrus L, Steinman R M. Lymphokine and non lymphokine mRNA levels in stimulated human T cells. J Exp Med 1986; 163: 922-37. 19 Kubo T, Towele C A, Markin H J, Treadwell B V. Stressinduced proteins in chondrocytes from patients with osteoarthritis. Ardhiis Rhewn 1985; 28: 1140-5. 20 Deguchi Y, Negoro S, Kishimoto S. Heat shock protein synthesis in peripheral blood mononuclear cells from patients with systemic lupus erythematosus. Biochem Biophys Res Commun 1987; 148: 1063-8.
