JHM Column Chen Chunhai and Zhou Zhou's team: The key role of SOX2-regulated astrocyte neurite plasticity in arsenic-induced metabolic disorders

Click on the blue words above to follow our first author: He Zhixin (Department of Labor Hygiene, Army Military Medical University) Corresponding author: Associate Professor Chen Chunhai (Department of Labor Hygiene, Army Military Medical University) Professor Zhou Zhou (School of Public Health, Zhejiang University) Corresponding Units: Department of Labor and Labor Hygiene, Army Medical University; School of Public Health, Zhejiang University; Brain Intelligence Research Center, Chongqing University School of Medicine Co-authored a research paper titled "SOX2 modulated astrocytic process plasticity is involved in inarsenic-induced metabolic disorders" in the Journal of Hazardous Materials (IF: 10.
6)
.

This study found that chronic arsenic exposure can reduce Glucose sensing in hypothalamic POMC neurons by impairing SOX2-regulated astrocytic process plasticity, thereby inducing metabolic disorders in mice.
The results provide a new central regulatory mechanism for revealing the occurrence of arsenic exposure-induced metabolic disorders in mice
.

Introduction Metabolic disorder diseases are clinically classified as a complex multi-disease cluster, including obesity, prediabetes, diabetes, and metabolic disorder syndromes
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In recent years, the occurrence of metabolic disorders induced by environmental factors has become more and more frequent.
As one of the harmful environmental pollutants, inorganic arsenic, its chronic exposure has been proved to be able to induce by interfering or impairing blood glucose metabolism, blood lipid metabolism and energy homeostasis.
Occurrence of metabolic disorders
.

However, the biological mechanism of metabolic disorders induced by arsenic exposure has not been fully elucidated, especially the related mechanism of central regulation of metabolism has not been reported in the literature.
It is of great significance for a comprehensive understanding of metabolic diseases caused by exposure to harmful environmental factors and the mechanism of astrocytes in regulating metabolic homeostasis
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Therefore, this study used the As exposure dose in the real environment of the population to establish a chronic As exposure mouse model, and assessed As exposure by detecting the series indicators of blood glucose homeostasis, energy metabolism indicators, and brain glucose absorption indicators in mice.
Effects on metabolic homeostasis and brain glucose sensitivity in mice
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Taking the plasticity of mouse hypothalamic astrocytes as an entry point, a mouse model of specifically knocking out SOX2 in astrocytes was constructed, combined with As exposure, to explore the role of SOX2 in antagonizing As-induced astrocytes.
The role and mechanism of impaired neurite plasticity and metabolic disorders in mice
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Figure 1: Chronic low-dose arsenic exposure induces disorder of blood glucose metabolism in mice As shown in Figure 1, chronic arsenic (As) exposure impaired glucose tolerance in mice and significantly increased fasting blood glucose levels
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In addition, it was found that As exposure significantly induced insulin resistance in mice
.

Then it was found that As exposure did not cause significant changes in fasting blood insulin levels in mice, and there was no significant difference in HOMA-β value, an indicator reflecting insulin secretion capacity, between As exposure group and control group
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However, the HOMA-IR value of As-exposed mice was significantly higher than that of control mice
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Subsequently, we further conducted insulin secretion experiments in mice.
The results were consistent with the above findings.
As exposure did not cause significant changes in the insulin secretion capacity of mice
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The above results indicate that chronic low-dose As exposure significantly impairs blood glucose homeostasis in mice and induces insulin resistance, but does not cause significant changes in insulin secretion levels
.

Figure 2: Chronic low-dose As exposure induces imbalance of energy homeostasis in mice To evaluate the impaired effect of chronic low-dose As exposure on energy homeostasis in mice, we used metabolic cages to control the energy homeostasis of mice after 14 weeks of exposure to As drinking water.
The mice underwent real-time monitoring of VO2, VCO2, RER and EE
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It was found that As exposure significantly reduced VO2, VCO2, RER, and EE in mice during the night cycle
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In addition, we also used a small animal infrared thermal imager to measure the body surface temperature of the interscapular back of mice, and found that As exposure could significantly reduce the temperature of the interscapular back skin of mice (Figure 2)
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Next, we detected the levels of TG and TC in the fasting serum of the two groups of mice, and found that the serum TG and TC levels of the As-exposed mice were significantly higher than those of the control mice
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The above results indicated that chronic low-dose As exposure significantly reduced the energy expenditure in mice and caused disturbances in the secretion levels of TG and TC hormones
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Figure 3: Chronic low-dose As exposure injures central glucose sensitivity in mice In order to explore the effect of As exposure on brain glucose sensitivity in mice, we injected 18F-FDG into two groups of mice intraperitoneally, and used small animal in vivo PET imaging technology to detect small animals.
Rat brain glucose uptake capacity was evaluated
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It was found that As exposure significantly reduced brain glucose uptake in mice
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In order to further explore the effect of As exposure on the glucose sensitivity of hypothalamic neurons, we performed immunofluorescence staining on the hypothalamus of neuronal cell activation marker c-FOS after two groups of mice were intraperitoneally injected with glucose for 2 h.
The results showed that As exposure Can significantly reduce the number of c-FOS positive cells in the hypothalamus VMH area, DMH area and ARC area
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Since POMC neurons are a type of neurons mainly involved in regulating energy metabolism homeostasis in the ARC region of the hypothalamus, in order to evaluate whether As exposure affects the glucose sensitivity of POMC neurons in the hypothalamus of mice, we injected glucose 2 into the mice.
After h, the mouse hypothalamus was double-labeled by immunofluorescence with POMC and c-FOS
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The results showed that As exposure could significantly reduce the number of c-FOS+POMC+ cells in the hypothalamus of mice
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In addition, we detected the expression of Pomc mRNA in the hypothalamus of the two groups of mice by real-time PCR technology, and found that As exposure could significantly reduce the expression of Pomc mRNA in the hypothalamus of mice (Figure 3)
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The above results indicated that chronic low-dose As exposure significantly impaired the glucose sensitivity of the mouse hypothalamus, especially decreased the glucose sensitivity of the hypothalamic POMC neurons
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Figure 4: Chronic low-dose As exposure disrupts hypothalamic astrocyte neurite plasticity.
Astrocytes play an important role in regulating central blood glucose supply, and astroglial neurite plasticity is important in maintaining blood-brain barrier function.
It plays a key role in permeability and neuron-astroglial glucose transport, so we will further explore the effect of As exposure on the plasticity of hypothalamic astrocyte protrusions
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First, we performed GFAP immunofluorescence staining on the hypothalamus of two groups of mice, and found that As exposure significantly reduced the total length of neurites and the number of neurite branches in astrocytes
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Further, we performed Golgi staining on the hypothalamus of the two groups of mice, and the results were consistent with previous findings that the As-exposed mice hypothalamic astrocytes had shorter neurite lengths and fewer neurite branches (Fig.
4).
)
.

Figure 5: Chronic low-dose As exposure reduces the encapsulation of blood vessels and POMC neurons by hypothalamic astrocytes.
Immunofluorescence labeling of thalamus with GFAP and vascular marker CD31
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It was found that As exposure significantly reduced the encapsulation of blood vessels by astrocytes
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To further explore the effect of As exposure on POMC neurons encapsulated by astrocytes, we performed immunofluorescence labeling of GFAP and POMC in mouse hypothalamus
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It was found that As exposure significantly reduced the encapsulation of POMC neurons by astrocytes (Fig.
5)
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We then confirmed this finding again by immunoelectron microscopy
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Taken together, the above results indicated that As exposure significantly disrupted the encapsulation of blood vessels and POMC neurons by astrocytes in the mouse hypothalamus, and resulted in impaired glucose sensitivity in the mouse hypothalamus
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Figure 6: Chronic low-dose As exposure inhibits SOX2 expression and activation of INSR/AKT pathway The expression of SOX2 in the hypothalamus of mice in the groups
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It was found that As exposure could significantly down-regulate the expression of SOX2
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Next, we examined the expression of INSR and the phosphorylation of AKT in mouse hypothalamus
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It was found that As exposure significantly inhibited the expression of INSR and the phosphorylation of AKT (Fig.
6)
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Taken together, the above results indicate that As exposure can significantly reduce the expression of SOX2 and inhibit the activation of the INSR/AKT pathway
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Figure 7: Astrocyte-specific knockout of SOX2 exacerbates As-induced metabolic disorders To further explore the role of SOX2 in As-induced metabolic disorders, we constructed a specific knockout of astrocytes hGfap-CreERT2 of SOX2;Sox2f/f mice
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We then used this SOX2-cKO mouse to study the effect of As exposure on metabolic homeostasis
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First, we found that SOX2-specific knockout alone did not significantly induce weight gain, glucose intolerance, and insulin resistance in mice
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However, SOX2-specific knockout worsened the effects of As exposure on body weight and random blood glucose in mice
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In addition, SOX2-specific knockdown also exacerbated As-induced glucose intolerance and insulin resistance (Fig.
7)
.

The above results suggest that the specific knockout of SOX2 in astrocytes can exacerbate As-induced metabolic disorders
.

Figure 8: Astrocyte-specific knockout of SOX2 exacerbates As-induced decline in glucose sensitivity, activation of insulin signaling, and impairment of astrocyte neurite plasticity.
Effects of hypothalamic POMC neurons on glucose sensitivity, we double-labeled POMC and c-FOS immunofluorescence in mouse hypothalamus
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It was found that knockdown of SOX2 in astrocytes alone did not significantly affect the number of c-FOS+POMC+ neurons in the hypothalamus, however, SOX2-specific knockout further reduced the number of As-induced c-FOS+POMC+ neurons.
drop
.

In addition, we also examined the effect of SOX2-specific knockout on As-induced plasticity damage in hypothalamic astrocytes by immunofluorescence technique, and found that SOX2-specific knockout further aggravated As-induced astrocytes There was a reduction in the number of cell branches and a reduction in the encapsulation of POMC neurons by astrocytes (Fig.
8)
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Furthermore, we examined the effect of SOX2 knockdown on As-induced inhibition of INSR/AKT pathway activation by Western Blot, and found that SOX2-specific knockout also worsened the As-induced inhibitory effect of INSR/AKT pathway
.

Figure 9: Specific overexpression of SOX2 in astrocytes can reverse the damage of astrocytic neurite plasticity caused by As exposure and the inhibition of INSR/AKT pathway to explore the role of SOX2 in As-induced primary astrocyte injury We performed GFAP immunofluorescence staining on primary astrocytes and found that specific knockout of SOX2 aggravated the As-induced damage of primary astrocyte neurite plasticity
.

In addition, we also found that specific knockdown of Sox2 also exacerbated As-induced inhibition of AKT phosphorylation and down-regulation of INSR expression (Fig.
9)
.

To further confirm the role of SOX2 in regulating astrocyte neurite plasticity and the INSR/AKT pathway, we constructed a lentivirus that specifically overexpressed SOX2 in astrocytes and used it to infect As-treated of primary astrocytes
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It was found that specific overexpression of astrocyte SOX2 could reverse As-induced astrocyte injury and the INSR/AKT pathway
.

Taken together, the above results suggest that astrocyte SOX2 plays an important role in As-induced astrocytic neurite plasticity and INST/AKT pathway (Fig.
9)
.

Summary This study aimed to investigate the key mechanism of SOX2-regulated astrocyte plasticity in arsenic-induced metabolic disorders in mice
.

The results show that specific knockout of SOX2 expression in astrocytes can exacerbate arsenic exposure-induced disturbance of glucose metabolism in mice, impaired plasticity of astrocyte neurites, decreased glucose sensitivity of hypothalamic POMC neurons, and INSR/AKT Signal blockade, and SOX2 overexpression reversed the toxic damage effects induced by arsenic exposure
.

In conclusion, the results of this study provide a new central regulatory mechanism and cellular target for the occurrence of metabolic disorders induced by chronic arsenic exposure, which will help to understand the role of SOX2-regulated astrocytic neurite plasticity in metabolic disorders induced by harmful environmental factors.
The role and role played in the occurrence of disease
.

About the author The first author: He Zhixin, a 2019 doctoral student in the Department of Labor Hygiene, Army Medical University, mainly engaged in the related research on the mechanism of heavy metal exposure-induced neurotoxicity effect, currently in Journal of Hazardous Materials, Ecotoxicology and Environmental Safety, Science of the Total Published 5 SCI papers in mainstream journals in the environmental field such as Environment
.

Corresponding author: Associate Professor Chen Chunhai, master tutor
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Mainly engaged in the study of neurotoxic effects induced by exposure to various environmental and occupational harmful factors such as heavy metals and electromagnetic radiation
.

A large number of studies have been carried out on the health hazards induced by a variety of environmental hazards
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He has published more than 20 SCI papers in high-level journals in related fields at home and abroad, such as Journal of Hazardous Materials, Science of the Total Environment, Cerebral Cortex, Stem Cells and Development, Ecotoxicology and Environmental Safety
.

Professor Zhou Zhou, doctoral supervisor
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National Excellent Youth, Chongqing Outstanding Youth
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Mainly engaged in basic research on the toxicological mechanism of heavy metal exposure and the induction of protection
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Innovative achievements have been made in the health hazard mechanism of exposure to various environmental heavy metal pollutants
.

He has published more than 80 SCI papers in high-level journals in related fields at home and abroad: Signal transduction and targeted therapy, Autophagy, Journal of Pineal Research, Journal of Hazardous Materials, Environmental International, Science of the Total Environment, Ecotoxicology and Environmental Safety
.

Contribution: Chen Chunhai and Zhou Zhou's team
.

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