Table 2.

Characteristics of the animal model/ Cell linestudies

Author Year, (Reference	animal model/ Cell line	Intervention/ duration	aim	Main Outcomes
Sassek et al. 2018 [33]	male Wistar rats/INS-1E cells and pancreatic islets	treated for 3 days with spexin	To identify the role, connections, and potential functions of spexin in endocrine pancreas.	Insulin secretion: - was reduced by spexin at 16 mM glucose level from cultured cells and isolated - Was decreased after injection with spexin in obese rats Spexin treatment: - increase in cultured cells and pancreatic islets cell viability and proliferation and proliferating cell nuclear antigen protein level. - decrease in insulin and Pdx gene expression.
Sassek et al. 2018 [33]	A pancreas obtained from a pig	three concentrations of glucose (1, 6, and 16 mM) and two periods of incubation (90 min and 24 h)	To identify the potential role of spexin in the physiology of the endocrine pancreas	1- presence of spexin inside the endocrine structures of the pancreas 2- glucose administration: - increase in spexin release from islets after a shortterm - decrease in spexin release after a long term - negative feedback loops between spexin andinsulin were found
Kolodziejski et al. 2018 [35]	murine 3 T3-L1 cell adipocytes and/or isolated human adipocytes.	treated with different concentrations of SPX (1.0, 10, 100, 1000 nM)	To investigate the effects of SPX on proliferation, cell viability, lipogenesis and lipolysis	SPX: - inhibits adipogenesis - down-regulates mRNA expression of pro-adipogenic genes such as PPARγ, C/EBPα, C/EBPβ and Fabp4. - Stimulates: lipolysis by increasing the phosphorylation of (HSL). - inhibits lipogenesis and glucose uptake in human adipocytes and murine 3 T3-L1 cells. - no effect on murine 3 T3-L1 cell proliferation and viability
Deng et al. 2018 [24]	spotted scat fish	fasting and refeeding periods(for 2 or 7 days)	To clarify the roles of Spx in the regulation of food-intake and reproduction in the spotted scat	A significant increase of: - spx expression in hypothalamus was observed after 2 and 7 days of food deprivation. A significant decrease of: - the spx transcript levels in the 7 day fasting group after refeeding 3 h after the scheduled feeding time
Wang et al. 2018 [25]	Tongue sole fish	- fasting and refeeding periods(for 2, 7, or 35 days) -tongue sole SPX14 (100 and 1000 ng/g BW dosage)	(1)to study the expression profiles of spexin mRNA in various tissues, (2) to evaluate the effects of feeding status on spexin mRNA levels, (3) to investigate the potential role of SPX in the control of appetite and reproduction of tongue sole	spexin mRNA: - could be detected in various tissues, notably in the brain. - Stimulated by fasting(the hypothalamic expression of SPX)
He et al. 2017	goldfish (hepatocytes and brain cell culture)	IP injection of D-(+)-glucose or human insulin	To study the Functional Link Between Food Intake and Spexin Expression	feeding could elevate: - plasma levels of glucose, insulin, and SPX with concurrent rises in insulin and SPX (mRNA) expression in the liver. Injection of glucose and insulin: - elevate in SPX mRNA level in the liver and brain areas involved in appetite control respectively insulin signal induced by glucose shows dual role in SPX regulation - (1) acting as an autocrine/paracrine signal to trigger SPX mRNA expression in the liver - (2) serving as an endocrine signal to induce SPX gene expression in the brain
Zheng et al. 2017 [27]	zebrafishspx1−/− mutant lines	Intracranial administration of SPX1	Spexin role in Food Intake in zebrafish tissue	the spx1−/− mutant fish: - higher food intake than the wild type (WT) fish. - Higher level of (AgRP1), - higher glucose, triacylglycerol and cholesterol in the serum than WT fish Intracranial administration of SPX1: - reduce the mRNA expression of the AgRP1.
Wu et al. 2016 [28]	Ya-fish (Schizothoraxprenanti)	Feeding- re feeding periods	identification, tissue distribution and mRNA expression of spexin responses to peri-prandial and fasting	SPX mRNA expression: - significantly increased in fed group than unfed groups after a meal, while the unfed group at 1 and 3 h substantially decreased than pre-prandial groups - significantly decreased during fasting for a week and sharply increased after refeeding on the 7th day, and then return to normal level on the 9th day.
Li et al. 2016 [10]	orange-spotted grouper fish (Epinepheluscoioides)	- IP administration of spexin-14(100 and 1000 ng/g BW) -Feeding- re feeding periods	Molecular cloning and functional characterization of spexininorange-spotted grouper	Tissue expression analysis: -spexin is highly expressed in the brain, liver and ovary Real time-PCR analysis: - up-regulation of hypothalamic expression of spexin by food deprivation. IP administration of spexin-14: -significantly elevated the mRNA levels of (POMC) - suppressed the orexin expression in the hypothalamus
Ge et al. 2016 [15]	Mouse Models of Fatty Liver Disease	IPadministrationof spexin (25 μg/kg) daily for 28–29 days	To examined the effects of spexin, on obesity, type 2 diabetes mellitus (T2DM), and HS in thigh fat diet induced obesity mice	spexin treatment cause: - weight loss in DIO mice - improved glucose tolerance - decrease insulin resistance and HbAlc - reduce: hepatic lipids by 60%, serum ALT and AST Incubation with spexin; - inhibited LCFA uptake by hepatocytes isolated from DIO mice - Spexin treatment
Walewsk et al. 2014 [7]	C57BL/6 J mice with DIO,Obese (DIO) adult female Wistarrat,omental and subcutaneous human fat samples	Spexin (35 mg/kg/day SC) in rats (25 mg/kg/day IP) in mice	Effects of Spexin on Adipocyte Uptake of Long Chain Fatty Acids and Weight in Rodents with Diet-Induced Obesity	Spexin injection in rats: - reduction in caloric intake 32% with corresponding weight loss - meal pattern was not affected Spexin injection in mice: - significantly reduction in the (RER) at night, and increased locomotion. Spexin incubation in vitro: - significantly inhibition of facilitated fatty acid (FA) uptake into DIO mouse adipocytes.
Wong et al. 2013 [3]	Goldfish (Carassiusauratus)	ICV injection of Goldfish SPX	To clarify the function of SPX as a satiety factor in gold fish	SPX mRNA levels: - elevated by food intake in the telencephalon, optic tectum, and hypothalamus brain injection of SPX: - inhibited both basal and NPY- or orexin induced feeding behavior and food consumption. - reduction in expression of NPY, AgRP, and apelin, - rises in CCK, CART, POMC, MCH, and CRH mRNA levels in brain areas

Abbreviation:Spx; Spexin,IP: Intraperitoneal, SC:subcutaneous, ICV: Intracerebroventricular, POMC;pro opiomelanocortin, DIO: Diet Induced Obesity, HSL: hormone sensitive lipase,AgRP1:agouti-relate protein 1, LCFA: long chain fatty acid,RER: respiratory exchange ratio CCK: cholecystokinin, CART: Cocaine-andamphetamine-regulated transcript, MCH: melanin-concentrating hormone, CRH: corticotropin-releasing hormone