RPL22 Overexpression Promotes Psoriasis-Like Lesion by Inducing Keratinocytes Abnormal Biological Behavior


doi: 10.3389/fimmu.2021.699900.


eCollection 2021.

Affiliations

Item in Clipboard

Jinrong Zeng et al.


Front Immunol.


.

Abstract


Background:

Keratinocytes of psoriasis have anti-apoptotic properties including delayed apoptosis process, accelerated proliferation metabolism and postponed differentiation process. However, the specific mechanism leading to the abnormal biological behavior of keratinocytes remains unclear.


Objectives:

We investigated the role of increased RPL22 expression in regulating the abnormal biological behavior of keratinocytes and the mechanism of regulation of RPL22 expression in skin lesions of psoriatic patients.


Methods:

We examined clinical samples and utilized cytokine-induced cell and IMQ-treated mouse models. We determined the expression and functions of RPL22 in vitro and in vivo.


Results:

We showed that RPL22 expression was significantly increased in the skin lesions of psoriasis patients and IMQ-treated psoriatic-like mice. Such increased expression is attributed to hyperacetylation of histone H3K27 in the promoter region of RPL22. Interestingly, overexpression of RPL22 enhanced keratinocyte proliferation by increasing cyclinD1 expression and accelerated CD4+T cells recruitment via upregulating CXCL10 expression. Finally, we demonstrated that RPL22 overexpression promoted psoriasiform phenotypes in IMQ-induced mouse skins.


Conclusions:

These findings suggested that RPL22 regulates keratinocytes abnormal biological behavior and contributes to the development of psoriatic phenotypes. Thus, RPL22 might be a novel potential molecular target for treatment of psoriasis.


Keywords:

CXCL10; CyclinD1; H3K27ac; RPL22; keratinocytes; psoriasis.

Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures


Figure 1



Figure 1

RPL22 expression is elevated in skin lesions of psoriasis patients. (A) The protein expression level of RPL22 in skin lesions derived from psoriasis patients and healthy controls. (B) The statistical analysis of the relative mRNA (left panel, n = 23) and protein expression levels(right panel, n = 7) of RPL22 in skin lesions derived from psoriasis patients and healthy controls. (C) IHC staining of RPL22 in skin lesions derived from psoriasis patients and healthy controls. (D) Correlation of human RPL22 mRNA expression in psoriatic skin with PASI scores (n = 23). Data represent the mean ± SEM. **P < 0.01, ***P < 0.001. Two-tailed unpaired Student’s t test was used.


Figure 2



Figure 2

RPL22 expression is elevated in skin lesions of IMQ mouse models. (A) The protein expression level of RPL22 in lesional skins from IMQ-exposed mice and vehicle-exposed mice. (B) The statistical analysis of the relative mRNA (left panel, n = 5) and protein expression levels (right panel, n = 3) of RPL22 in lesional skins from IMQ-exposed mice and vehicle-exposed mice. (C) IHC staining of RPL22 in lesional skins from IMQ-exposed mice vehicle-exposed mice. (D) The mRNA expression level of RPL22 in psoriatic-like cells model on HaCaT cells stimulating with a cocktail of cytokines including Oncostatin-M, IL-1α, IL-17A, IL-22, and TNF-α(M5). Data (D) are representative of at least 3 independent experiments. Data represent the mean ± SEM. *P < 0.05. Two-tailed unpaired Student’s t test was used.


Figure 3



Figure 3

RPL22 promotes the proliferation of HaCaT cells and inhibits apoptosis of KCs. (A) Selection and verification of RPL22 siRNA intervention efficiency in HaCaT cells (n = 3). (B) Overexpression efficiency verification of RPL22 overexpressed plasmid (OE-RPL22) in HaCaT cells (n = 3). (C) Microscopy to observe cell proliferation state in HaCaT cells at 72hours. (D) CCK-8 assay to detect the cell proliferation level in HaCaT cells transfected with si-RPL22 or OE-RPL22 and their controls. (E) Flow cytometry to detect the apoptosis of KCs transfected with si-RPL22 or OE-RPL22 and their controls (n = 3). (F) Statistical analysis data to show the difference of four groups at 48 hours. All data are representative of at least 3 independent experiments. Data represent the mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. NS, not significant. Two-tailed unpaired Student’s t test was used.


Figure 4



Figure 4

RPL22 is involved in the cell cycle. (A) Flow cytometry to detect the cell cycle of HaCaT cells transfected with si-RPL22 or OE-RPL22 and their controls (n = 3). (B) Statistical analysis data to show the difference of four groups at 48 hours. (C) The mRNA levels of cell cycle-related protein including CyclinD1, CyclinA2 and CDK2 in HaCaT cells transfected with si-RPL22 or OE-RPL22 and their controls (n = 3). All data are representative of at least 3 independent experiments. Data represent the mean ± SEM. *P < 0.05, **P < 0.01. NS, not significant. Two-tailed unpaired Student’s t test was used.


Figure 5



Figure 5

RPL22 promotes cell chemotaxis. (A) Chemotaxis assay to detect the chemotaxis efficiency of KCs transfected with si-RPL22 or OE-RPL22 and their controls on CD4+T cells (n = 3). (B) Cell count to statistically analyze the chemotaxis differences (n = 3). (C) RT-qPCR to detect the mRNA levels of chemokines of KCs transfected with si-RPL22 or OE-RPL22 and their controls including CXCL10, CCL5 and CCL20 (n=3). (D) RT-qPCR to detect the mRNA levels of psoriasis-associated cytokines of KCs transfected with si-RPL22 or OE-RPL22 and their controls including IL-1β, IL-23, TNF-α and IL-17a (n = 3). All data are representative of at least 3 independent experiments. Data represent the mean ± SEM. *P < 0.05, **P < 0.01. NS, not significant. Two-tailed unpaired Student’s t test was used.


Figure 6



Figure 6

The hyperacetylation level in the promoter region of RPL22 elevates its expression. (A) Electrophoretic image of ChIP assay result showed the histone H3K27 acetylation level on RPL22 promoter in skin lesions derived from PV patients was significantly higher than healthy controls (n = 3). (B) ChIP-qPCR showed the enrichment of H3K27ac in RPL22 promoter in all three skin lesions derived from PV patients was significantly higher than healthy controls.


Figure 7



Figure 7

RPL22 overexpression accelerates the development of psoriatic lesions (A) Clinical efficacies on skin lesions in IMQ mice models after RPL22 intervention in four groups of si-RPL22, si-NC, OE-RPL22 and OE-NC. (B) H&E staining of lesional skin from mice in the above four groups to show pathological manifestation including acanthosis and dermal inflammatory cells infiltration. (C) RT-qPCR to detect the RPL22 mRNA expression level in lesional skin from IMQ-induced psoriasis-like mouse model intradermally injected with si-RPL22 or si-NC or OE-RPL22 or OE-NC (n = 5). (D) Acanthosis was quantified for IMQ-induced mice treated with si-RPL22 or si-NC or OE-RPL22 or OE-NC (n = 5). Data represent the mean ± SEM. *P < 0.05, **P < 0.01. Two-tailed unpaired Student’s t test was used.


Figure 8



Figure 8

Schematic illustration of RPL22 contributing to the pathogenesis of psoriasis. In psoriasis patients, hyperacetylation of histone H3K27 in the promoter region of RPL22 in the skin lesions of psoriasis patients induces RPL22 overexpression, overexpressed RPL22 up-regulates CyclinD1 expression to promote the KCs proliferation and enhancing CXCL10 expression to induce CD4+ T cells chemotaxis, Both of them secrete cytokines including IL-23, IL-6, IL-1β and IL-17a to aggravate the occurrence and development of psoriasis.

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