Limits...
Lipoic acid plays a role in scleroderma: insights obtained from scleroderma dermal fibroblasts.

Tsou PS, Balogh B, Pinney AJ, Zakhem G, Lozier A, Amin MA, Stinson WA, Schiopu E, Khanna D, Fox DA, Koch AE - Arthritis Res. Ther. (2014)

Bottom Line: In contrast, TIMP-1 levels were higher in SSc but DHLA had minimal effect.In general, DHLA showed better efficacy than NAC in most cases.Our study suggests that thiol antioxidants, including NAC and LA/DHLA, could be beneficial for patients with SSc.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine, Division of Rheumatology, University of Michigan Medical School, 109 Zina Pitcher Dr., 4388 BSRB, Ann Arbor, MI 48109-2200, USA. ptsou@umich.edu

ABSTRACT

Introduction: Systemic sclerosis (SSc) is a connective tissue disease characterized by fibrosis of the skin and organs. Increase in oxidative stress and platelet-derived growth factor receptor (PDGFR) activation promote collagen I (Col I) production, leading to fibrosis in SSc. Lipoic acid (LA) and its active metabolite dihydrolipoic acid (DHLA) are naturally occurring thiols that act as cofactors and antioxidants, and are produced by lipoic acid synthetase (LIAS). The goal of this study was to examine whether LA and LIAS was deficient in SSc patients and determine the effect of DHLA on the phenotype of SSc dermal fibroblasts. N-acetylcysteine (NAC), a commonly used thiol antioxidant, was included as a comparison.

Methods: Dermal fibroblasts were isolated from healthy subjects and patients with diffuse cutaneous SSc. Matrix metalloproteinase (MMPs), tissue inhibitors of MMPs (TIMP), plasminogen activator inhibitor-1 (PAI-1) and LIAS were measured by ELISA. The expression of Col I was measured by immunofluorescence, hydroxyproline assay, and quantitative PCR. PDGFR phosphorylation and α-smooth muscle actin (α-SMA) was measured by Western blotting. Student's t-tests were performed for statistical analysis and p-values of less than 0.05 with two-tailed analysis were considered statistically significant.

Results: The expression of LA and LIAS in SSc dermal fibroblasts was lower than normal fibroblasts, however LIAS was significantly higher in SSc plasma and appeared to be released from monocytes. DHLA lowered cellular oxidative stress, and decreased PDGFR phosphorylation, Col I, PAI-1, and α-SMA expression in SSc dermal fibroblasts. It also restored the activities of phosphatases that inactivated the PDGFR. SSc fibroblasts produced lower levels of MMP-1 and 3, and DHLA increased them. In contrast, TIMP-1 levels were higher in SSc but DHLA had minimal effect. Both DHLA and NAC increased MMP-1 activity when SSc cells were stimulated with PDGF. In general, DHLA showed better efficacy than NAC in most cases.

Conclusions: DHLA not only acts as an antioxidant but also an antifibrotic since it has the ability to reverse the profibrotic phenotype of SSc dermal fibroblasts. Our study suggests that thiol antioxidants, including NAC and LA/DHLA, could be beneficial for patients with SSc.

Show MeSH

Related in: MedlinePlus

The effect of dihydrolipoic acid on the platelet-derived growth factor receptor pathway and phosphatases. (A) Phosphorylated platelet-derived growth factor receptor (p-PDGFR) after PDGF stimulation at various time points (n = 3 NL and SSc subjects). NS, Nonstimulated. Enzymatic activities of protein tyrosine phosphatase 1B (PTP1B) (B), Src homology 2 domain–containing protein tyrosine phosphatase 2 (SHP-2) (C) and density-enhanced phosphatase 1 (DEP-1) (D) in fibroblasts with or without dihydrolipoic acid (DHLA) or N-acetylcysteine (NAC). Results are expressed as mean ± SEM. P < 0.05 was considered significant.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4558991&req=5

Fig2: The effect of dihydrolipoic acid on the platelet-derived growth factor receptor pathway and phosphatases. (A) Phosphorylated platelet-derived growth factor receptor (p-PDGFR) after PDGF stimulation at various time points (n = 3 NL and SSc subjects). NS, Nonstimulated. Enzymatic activities of protein tyrosine phosphatase 1B (PTP1B) (B), Src homology 2 domain–containing protein tyrosine phosphatase 2 (SHP-2) (C) and density-enhanced phosphatase 1 (DEP-1) (D) in fibroblasts with or without dihydrolipoic acid (DHLA) or N-acetylcysteine (NAC). Results are expressed as mean ± SEM. P < 0.05 was considered significant.

Mentions: Because LA was lower in SSc dermal fibroblasts and relieved oxidative stress, we hypothesized that adding DHLA, its active metabolite, to the cells would change their profibrotic phenotype back to normal. We first examined the effect of DHLA on PDGFR activation. In NL dermal fibroblasts, PDGF-stimulated PDGFR phosphorylation (p-PDGFR) was maximal at 30 minutes and decreased significantly at 1 hour (Figure 2A). In contrast, p-PDGFR maximized at 10 minutes and remained phosphorylated at 1 hour in SSc dermal fibroblasts. In the presence of DHLA, p-PDGFR peaked at 10 and 30 minutes for NL fibroblasts. However, PDGF did not induce p-PDGFR in the presence of DHLA in SSc fibroblasts. The time course of p-PDGFR with or without DHLA significantly decreased the extent of p-PDGFR in both NL and SSc. This suggests that the excessive p-PDGFR seen in SSc dermal fibroblasts is due to increased oxidative stress, as a thiol antioxidant could reduce it.Figure 2


Lipoic acid plays a role in scleroderma: insights obtained from scleroderma dermal fibroblasts.

Tsou PS, Balogh B, Pinney AJ, Zakhem G, Lozier A, Amin MA, Stinson WA, Schiopu E, Khanna D, Fox DA, Koch AE - Arthritis Res. Ther. (2014)

The effect of dihydrolipoic acid on the platelet-derived growth factor receptor pathway and phosphatases. (A) Phosphorylated platelet-derived growth factor receptor (p-PDGFR) after PDGF stimulation at various time points (n = 3 NL and SSc subjects). NS, Nonstimulated. Enzymatic activities of protein tyrosine phosphatase 1B (PTP1B) (B), Src homology 2 domain–containing protein tyrosine phosphatase 2 (SHP-2) (C) and density-enhanced phosphatase 1 (DEP-1) (D) in fibroblasts with or without dihydrolipoic acid (DHLA) or N-acetylcysteine (NAC). Results are expressed as mean ± SEM. P < 0.05 was considered significant.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4558991&req=5

Fig2: The effect of dihydrolipoic acid on the platelet-derived growth factor receptor pathway and phosphatases. (A) Phosphorylated platelet-derived growth factor receptor (p-PDGFR) after PDGF stimulation at various time points (n = 3 NL and SSc subjects). NS, Nonstimulated. Enzymatic activities of protein tyrosine phosphatase 1B (PTP1B) (B), Src homology 2 domain–containing protein tyrosine phosphatase 2 (SHP-2) (C) and density-enhanced phosphatase 1 (DEP-1) (D) in fibroblasts with or without dihydrolipoic acid (DHLA) or N-acetylcysteine (NAC). Results are expressed as mean ± SEM. P < 0.05 was considered significant.
Mentions: Because LA was lower in SSc dermal fibroblasts and relieved oxidative stress, we hypothesized that adding DHLA, its active metabolite, to the cells would change their profibrotic phenotype back to normal. We first examined the effect of DHLA on PDGFR activation. In NL dermal fibroblasts, PDGF-stimulated PDGFR phosphorylation (p-PDGFR) was maximal at 30 minutes and decreased significantly at 1 hour (Figure 2A). In contrast, p-PDGFR maximized at 10 minutes and remained phosphorylated at 1 hour in SSc dermal fibroblasts. In the presence of DHLA, p-PDGFR peaked at 10 and 30 minutes for NL fibroblasts. However, PDGF did not induce p-PDGFR in the presence of DHLA in SSc fibroblasts. The time course of p-PDGFR with or without DHLA significantly decreased the extent of p-PDGFR in both NL and SSc. This suggests that the excessive p-PDGFR seen in SSc dermal fibroblasts is due to increased oxidative stress, as a thiol antioxidant could reduce it.Figure 2

Bottom Line: In contrast, TIMP-1 levels were higher in SSc but DHLA had minimal effect.In general, DHLA showed better efficacy than NAC in most cases.Our study suggests that thiol antioxidants, including NAC and LA/DHLA, could be beneficial for patients with SSc.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine, Division of Rheumatology, University of Michigan Medical School, 109 Zina Pitcher Dr., 4388 BSRB, Ann Arbor, MI 48109-2200, USA. ptsou@umich.edu

ABSTRACT

Introduction: Systemic sclerosis (SSc) is a connective tissue disease characterized by fibrosis of the skin and organs. Increase in oxidative stress and platelet-derived growth factor receptor (PDGFR) activation promote collagen I (Col I) production, leading to fibrosis in SSc. Lipoic acid (LA) and its active metabolite dihydrolipoic acid (DHLA) are naturally occurring thiols that act as cofactors and antioxidants, and are produced by lipoic acid synthetase (LIAS). The goal of this study was to examine whether LA and LIAS was deficient in SSc patients and determine the effect of DHLA on the phenotype of SSc dermal fibroblasts. N-acetylcysteine (NAC), a commonly used thiol antioxidant, was included as a comparison.

Methods: Dermal fibroblasts were isolated from healthy subjects and patients with diffuse cutaneous SSc. Matrix metalloproteinase (MMPs), tissue inhibitors of MMPs (TIMP), plasminogen activator inhibitor-1 (PAI-1) and LIAS were measured by ELISA. The expression of Col I was measured by immunofluorescence, hydroxyproline assay, and quantitative PCR. PDGFR phosphorylation and α-smooth muscle actin (α-SMA) was measured by Western blotting. Student's t-tests were performed for statistical analysis and p-values of less than 0.05 with two-tailed analysis were considered statistically significant.

Results: The expression of LA and LIAS in SSc dermal fibroblasts was lower than normal fibroblasts, however LIAS was significantly higher in SSc plasma and appeared to be released from monocytes. DHLA lowered cellular oxidative stress, and decreased PDGFR phosphorylation, Col I, PAI-1, and α-SMA expression in SSc dermal fibroblasts. It also restored the activities of phosphatases that inactivated the PDGFR. SSc fibroblasts produced lower levels of MMP-1 and 3, and DHLA increased them. In contrast, TIMP-1 levels were higher in SSc but DHLA had minimal effect. Both DHLA and NAC increased MMP-1 activity when SSc cells were stimulated with PDGF. In general, DHLA showed better efficacy than NAC in most cases.

Conclusions: DHLA not only acts as an antioxidant but also an antifibrotic since it has the ability to reverse the profibrotic phenotype of SSc dermal fibroblasts. Our study suggests that thiol antioxidants, including NAC and LA/DHLA, could be beneficial for patients with SSc.

Show MeSH
Related in: MedlinePlus