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Comment on Azmahani et al. “Steroidogenic enzymes, their related transcription factors and nuclear receptors in human sebaceous glands under normal and pathological conditions”
Journal of Steroid Biochemistry & Molecular Biology 155 (2016) 177
Contents lists available at ScienceDirect
Journal of Steroid Biochemistry & Molecular Biology journal homepage: www.elsevier.com/locate/jsbmb
Letter to the Editor Comment on Azmahani et al. “Steroidogenic enzymes, their related transcription factors and nuclear receptors in human sebaceous glands under normal and pathological conditions”
Dear Editor Adrenocortical cancer is one of the “areas of interest” of our scientific group, therefore, we have approached with interest the article published by Azmahani et al. [1] on Journal of Steroid Biochemistry & Molecular Biology. In this study the authors aimed to investigate the status of steroidogenic enzymes, sex steroid receptors and transcription factors in 22 human normal sebaceous glands, 12 specimens of sebaceous nevus, 12 of sebaceous hyperplasia, 3 of sebaceoma and 10 of sebaceous carcinoma to explore their possible roles in in situ steroid production in human skin. The localization of steroidogenic enzymes, sex steroid receptors and transcription factors in the human sebaceous glands under normal and pathological conditions has been investigated by immunohistochemistry (IHC). Moreover, to better understand the underlying pathways and regulation of steroidogenic enzymes in the sebaceous gland, the expression of 3b-HSD1, CYP11A1, StAR,17b-HSD5, 5a-red1, NGFI-B, SF-1 and GATA6 mRNA has been detected by RT-qPCR in both SZ95 sebocytes, with or without forskolin, and in H295R cells, an adrenal carcinoma cell line used as positive control. The results of IHC, which were described with very clear and nice pictures, demonstrated that the protein expression of 3bHSD1, CYP11A1, StAR, 17b-HSD5, CYP17A1, 5a-red1, PRB, AR and NGFI-B was higher in human normal sebaceous glands compared to the pathological sebaceous glands. Using IHC, the authors failed to demonstrate steroidogenic factor 1 (SF-1) expression in all investigated tissues. Intriguingly, SF-1 mRNA expression was significantly higher in SZ95 sebocytes, compared to H295R adrenocortical cells, and, in the text, the authors speculated about the meaning of the difference between SF-1 mRNA and protein expression. During the study of this scientific paper, I realized that the SF1 primers sequences reported by the authors were not correspondent to steroidogenic factor 1. The choice of the appropriate primers is probably the most important factor affecting the
http://dx.doi.org/10.1016/j.jsbmb.2015.09.031 0960-0760/ ã 2015 Elsevier Ltd. All rights reserved.
polymerase chain reaction (PCR) [2]. In the text, the authors were referring to SF-1 as steroidogenic factor 1, but a simple checking of the primers by using a software tool like Primer Blast from NBCI, allowed me to verify that the primers used in the paper are absolutely not specific for the steroidogenic factor 1 gene, but they amplify another gene: the splicing factor 1. Therefore, a possible explanation regarding the significant difference in the mRNA expression levels of SF-1 in SZ95 sebocytes, compared to H295R, and the different expression of SF-1 between human sebaceous glands (SF-1 at protein level) and SZ95 sebocytes (SF-1 at messenger level) could be the improper choice of primers set. This evidence implies that part of this study need to be reassessed in order to have definitive conclusion on the expression of steroidogenic factor-1. References [1] A. Azmahani, Y. Nakamura, S.J.A. Felizola, Y. Ozawa, K. Ise, T. Inoue, K.M. McNamara, M. Doi, H. Okamura, C.C. Zouboulis, S. Aiba, H. Sasano, Steroidogenic enzymes, their related transcription factors and nuclear receptors in human sebaceous glands under normal and pathological conditions, J. Steroid Biochem. 144 (2014) 268–279. [2] J. Ye, G. Coulouris, I. Zaretskaya, I. Cutcutache, S. Rozen, T.L. Madden, PrimerBLAST: a tool to design target-specific primers for polymerase chain reaction, BMC Bioinf. 13 (2012) 134.
Claudia Pivonello Maria Cristina De Martino Annamaria Colao Rosario Pivonello* Dipartimento di Medicina Clinica e Chirurgia, Università di Napoli “Federico II”, Italy * Corresponding author at: Dipartimento di Medicina Clinica e Chirurgia, “Federico II” University, Via Sergio Pansini, 5, 80131 Naples, Italy. Fax: +39 0817464983. E-mail address: [email protected] (R. Pivonello). Received 8 July 2015 Accepted 18 September 2015 Available online 28 September 2015
Contents lists available at ScienceDirect
Journal of Steroid Biochemistry & Molecular Biology journal homepage: www.elsevier.com/locate/jsbmb
Letter to the Editor Comment on Azmahani et al. “Steroidogenic enzymes, their related transcription factors and nuclear receptors in human sebaceous glands under normal and pathological conditions”
Dear Editor Adrenocortical cancer is one of the “areas of interest” of our scientific group, therefore, we have approached with interest the article published by Azmahani et al. [1] on Journal of Steroid Biochemistry & Molecular Biology. In this study the authors aimed to investigate the status of steroidogenic enzymes, sex steroid receptors and transcription factors in 22 human normal sebaceous glands, 12 specimens of sebaceous nevus, 12 of sebaceous hyperplasia, 3 of sebaceoma and 10 of sebaceous carcinoma to explore their possible roles in in situ steroid production in human skin. The localization of steroidogenic enzymes, sex steroid receptors and transcription factors in the human sebaceous glands under normal and pathological conditions has been investigated by immunohistochemistry (IHC). Moreover, to better understand the underlying pathways and regulation of steroidogenic enzymes in the sebaceous gland, the expression of 3b-HSD1, CYP11A1, StAR,17b-HSD5, 5a-red1, NGFI-B, SF-1 and GATA6 mRNA has been detected by RT-qPCR in both SZ95 sebocytes, with or without forskolin, and in H295R cells, an adrenal carcinoma cell line used as positive control. The results of IHC, which were described with very clear and nice pictures, demonstrated that the protein expression of 3bHSD1, CYP11A1, StAR, 17b-HSD5, CYP17A1, 5a-red1, PRB, AR and NGFI-B was higher in human normal sebaceous glands compared to the pathological sebaceous glands. Using IHC, the authors failed to demonstrate steroidogenic factor 1 (SF-1) expression in all investigated tissues. Intriguingly, SF-1 mRNA expression was significantly higher in SZ95 sebocytes, compared to H295R adrenocortical cells, and, in the text, the authors speculated about the meaning of the difference between SF-1 mRNA and protein expression. During the study of this scientific paper, I realized that the SF1 primers sequences reported by the authors were not correspondent to steroidogenic factor 1. The choice of the appropriate primers is probably the most important factor affecting the
http://dx.doi.org/10.1016/j.jsbmb.2015.09.031 0960-0760/ ã 2015 Elsevier Ltd. All rights reserved.
polymerase chain reaction (PCR) [2]. In the text, the authors were referring to SF-1 as steroidogenic factor 1, but a simple checking of the primers by using a software tool like Primer Blast from NBCI, allowed me to verify that the primers used in the paper are absolutely not specific for the steroidogenic factor 1 gene, but they amplify another gene: the splicing factor 1. Therefore, a possible explanation regarding the significant difference in the mRNA expression levels of SF-1 in SZ95 sebocytes, compared to H295R, and the different expression of SF-1 between human sebaceous glands (SF-1 at protein level) and SZ95 sebocytes (SF-1 at messenger level) could be the improper choice of primers set. This evidence implies that part of this study need to be reassessed in order to have definitive conclusion on the expression of steroidogenic factor-1. References [1] A. Azmahani, Y. Nakamura, S.J.A. Felizola, Y. Ozawa, K. Ise, T. Inoue, K.M. McNamara, M. Doi, H. Okamura, C.C. Zouboulis, S. Aiba, H. Sasano, Steroidogenic enzymes, their related transcription factors and nuclear receptors in human sebaceous glands under normal and pathological conditions, J. Steroid Biochem. 144 (2014) 268–279. [2] J. Ye, G. Coulouris, I. Zaretskaya, I. Cutcutache, S. Rozen, T.L. Madden, PrimerBLAST: a tool to design target-specific primers for polymerase chain reaction, BMC Bioinf. 13 (2012) 134.
Claudia Pivonello Maria Cristina De Martino Annamaria Colao Rosario Pivonello* Dipartimento di Medicina Clinica e Chirurgia, Università di Napoli “Federico II”, Italy * Corresponding author at: Dipartimento di Medicina Clinica e Chirurgia, “Federico II” University, Via Sergio Pansini, 5, 80131 Naples, Italy. Fax: +39 0817464983. E-mail address: [email protected] (R. Pivonello). Received 8 July 2015 Accepted 18 September 2015 Available online 28 September 2015