J Food Bioact

Journal of Food Bioactives, ISSN 2637-8752 print, 2637-8779 online

Journal website www.isnff-jfb.com

Review

Volume 31, September 2025, pages 63-75

An update on therapeutic potential of earthworm extract

Figure 1. Mechanisms by which EE alleviates pulmonary fibrosis and liver fibrosis. (a) EE inhibits Bax expression, promoted Bcl2 expression, alters mitochondrial membrane potential, and inhibits cytochrome c release and Caspase-3/9 expression. (b) EE promotes Nrf2 nuclear translocation, promoted downstream NQO-1/HO-1 expression, and inhibits ROS levels. (c) EE inhibits the expression of TGF-β1, α-SMA, PAI-1, IL-1β and promoted the expression of E-cadherin.

Figure 2. The mechanisms by which EE promotes apoptosis of cancer cells. (a) EE inhibits wnt/β-catenin pathway. (b) EE alters mitochondrial membrane permeability and releases cytochrome c. (c) EE promotes the expression of caspase3/4/5/10 in the cells.

**Table 1.** Bioactivity of EE and mechanism
Sample	Source/Variety	In vitro/in vivo	Model	Dose/time of duration	mechanism	target/pathway	ref
Notes: “–” represents the source or dose was not shown. For lumbrokinase, we show the source instead of variety.
Lumbrokinase	Canada RNA biochemical Inc., Richmond, BC, Canada	In vivo	I-R injury in male Sprague-Dawley rats	1,10 μg/kg	Inhibited TLR4 expression, the phosphorylation of JNK, NF-κB, IκBα and the protein expression of COX-2, iNOS and MMP-9	TLR4, JNK, NF-Κb, IκBα, COX-2, iNOS, MMP9	(Wang et al., 2016)
Lumbrokinase	Canada RNA biochemical Inc., Richmond, BC, Canada	In vivo	mice	0.1, 1 mg/kg/7 days	Decreased the expression of IRE1 and inhibited the expression of transcription factors, XBP-1, caspase-12 and NF-κB, reduced NLRP3 inflammasome	IRE1, XBP-1, NF-κB, NLRP3, caspase-12	(Wang et al., 2022)
Earthworm active protein	Xian Guanmao Biotechnology Co., Ltd.	In vivo	Hyperlipidemic SD rats	50, 100, 200 mg/kg/28 days	Increased lipase activity and enhanced intestinal cholesterol metabolism transformation	–	(Yuan et al., 2018)
Polysaccharide-protein complex (PPC-DV)	Dendrobaena veneta	In vitro	Blood from peripheral vein	25, 50, 100 μg/ml	Targeted the P2Y12 receptor, COX-1 and PAR-1 pathways	P2Y12R, COX-1, PAR-1	(Poniedzialek et al., 2022)
Dilong extract	Pheretima vulgaris	In vivo and vitro	Collagen-induced arthritis in mice and murine RAW 264.7 macrophages	5, 10 g/kg/28 days and 75, 150, 300 μg/kg/1 days	Inhibited the NF-κB p65 activation, regulated the ratio of Th1/Th2 cells and improved the abnormal levels of inflammatory cytokines	NF-κB p65	(Bao et al., 2022)
Earthworm extract	El-Bagour, Menoufia, Egypt	In vivo	Acrylamide induced reproductive Injury in male rats	300 mg/ml/15 days	Inhibited the expression of P53 and Ki67, restored testicular antioxidant balance	P53, Ki67	(Ahmed et al., 2022)
Lumbrokinase	–	in vivo	I-R induced testicular injury in rats	80mg/kg	Inhibited the expression of Bax	Bax	(Danarto et al., 2019)
Earthworm extract	–	In vivo	Mouse wound model	0.1 ml	Reduced the ill-effects of inflammation, accelerated the secretion of hydroxyproline and TGF-β, increased the synthesis of collagen, and promoted the proliferation of blood capillary, and fibroblast. Accelerated the removal of dead tissue and foreign bodies by accelerating the production of IL- 6, white blood cells and platelets	TGF-β, IL- 6	(Deng et al., 2018)
Earthworm active ingredients	Eisenia foetida	In vitro	Tunicamycin induced L-02 cells apoptosis	0.1, 0.2, 0.4 mg/L/24 hours	Inhibited mRNA and protein expressions of GRP78, PERK, ATF4, Elf2α, CHOP, Bax, relieved ERS, he protein and mRNA expressions of Bcl-2, promoted the proliferation of L-02 cells.	GRP78, PERK, ATF4, Elf2α, CHOP, Bax, ERS, Bcl-2	(Duan et al., 2018)
Earthworm extract	Allolobophora caliginosa	In vivo	SiNPs induced liver injury in male albino rats	500, 1,000 mg/kg/30 days	Regulated the antioxidant balance of the liver	–	(Sadek et al., 2016)
Earthworm coelomic fluid	Allolobophora caliginosa	In vivo	Gentamicin induced hepatorenal toxicity in rats	45 mg/kg/7 days	Regulated the balance of oxidative system in the body	–	(Mohamed et al., 2020)
Earthworm extract	Perionyx excavates	In vivo	HFD-induced non-alcoholic fatty liver in guinea pigs	0.3, 1.4, 6.8 μg/kg	Inhibited the expression of TC, TG and LDL-C, alleviated liver injury in guinea pigs	TC, TG, LDL-C	(Deng et al., 2021)
Earthworm extract	–	In vivo	Silica-induced pulmonary fibrosis in mice	600 U/7, 14, 28 days	Activated Nrf2 pathway, inhibited oxidative stress, alleviated silica-induced apoptosis of lung epithelial cells and EMT	Nrf2 pathway, EMT pathway	(Yang et al., 2016)
Dilong extract	Guang dilong	In vivo	Bleomycin-induced pulmonary fibrosis mice	1.175, 2.35, 4.7g/kg/14 days	Combined with TGF-β1, inhibited the expression of α-SAM	α-SAM	(Wang et al., 2019)
Pheretima protein P2	Guang dilong	In vitro and vivo	TGF-β induced abnormal proliferation in MRC5 cells and bleomycin induced pulmonary fibrosis in mice	13.125μg/ml/2h and 5mg/kg/22days	Inhibited the protein expression of smad2/3 and p-smad2/3, inhibited the expression of α-SMA, Vimentin and Collagen I and increased the expression of E-cadherin	smad2/3, p-smad2/3, α-SMA, Vimentin, Collagen I, E-cadherin	(Li et al., 2022)
Water extract of earthworms	Pheretima aspergillum	In vivo and vitro	CCL4- induced liver fibrosis in mouse and TGFβ1 treated LX-2 HSCs and AML-12 cells	100,200 mg/kg/7 days and 30, 100 μg/ml/24 hours	Enhanced AMPK/GSK3β/Nrf2 cascade by activating LKB1, inhibited HSC activation, hepatocyte apoptosis and ferroptosis	LKB1, AMPK/GSK3β/Nrf2 cascade	(Zhang et al., 2024)
Earthworm granulation tissue extract	Eisenia fetida	In vivo	STZ induced diabetic rats	50, 70%/21 days	Scavenged free radicals, inhibited inflammatory pathways, reduced the levels of TNF-α, IL-1β, LPO, IL-6, promoted the synthesis of connective tissue framework protein and the expression of growth promoters	TNF-α, IL-1β, LPO, IL-6	(Elkhalifa et al., 2024)
Earthworm extract	–	In vivo	burn wound model on skin of mice	0.1 ml/3, 7, 11, 15 days	decreased edema, suppressed fibrosis, activated angiogenesis and epithelial regeneration, inhibited scar formation, and reduced the risk of infection.	–	(He et al., 2021)
Earthworm extract	Pheretima aspergillum	In vitro	Osteoblast-like MG-63 cells and RAW 264.7 macrophage cells	500 ng/ml – 12 mg/ml/2 days	increased osteoblast proliferation and differentiation, matrix calcium deposition and the expression levels of alkaline phosphatase (ALP), osteopontin (OPN) and osteocalcin (OCN), reduced the tartrate-resistant acid phosphatase (TRAP) activity of osteoclasts	ALP, OPN, OCN, TRAP	(Fu et al., 2014)
Earthworm active protein	–	In vitro	NIH3T4 cell line	37.5, 75, 150 mg/ml	Activated the MEK/ERK signaling pathway, promoted the cell cycle from G1 phase to S phase, and promoted cell proliferation	MEK/ERK signaling pathway	(Song et al., 2016)
Earthworm extract	–	In vitro	Human ovarian cancer cells A2780	400, 800 mg/ml/72 hours	Inhibited wnt / βcatenin pathway and promoted apoptosis	wnt / βcatenin pathway	(Chen et al., 2023)
Earthworm powder	Lumbricus terrestris	In vitro	Breast cancer cells (MCF-7) and prostate cancer cells (PC-3)	100, 200, 400 μg/ml/2 4 hours	Changed the permeability of the mitochondrial membrane and released cytochrome c	–	(Shafi and Faleh, 2019)
Earthworm extract	Perionyx excavates	In vivo	S180 tumor-bearing mice	30, 60, 90 mg/kg/25 days	Reduced the level of LDH, regulated the expression of Bax and Bcl2 protein	LDH, Bax, Bcl2	(Deng et al., 2019)
Earthworm coelomic fluid	Dendrobaena veneta	In vitro	A549 cells	125, 187.5, 250 μg/ml/24 hours	Increased expression of caspase3/4/5/10	caspase3/4/5/10	(Fiolka et al., 2019)
Protein-carbohydrate fraction (PE)	Dendrobaena veneta	In vitro	HT-29 cells	10–200 μg/ml	Inhibited mitochondrial metabolism, up-regulated caspase-3 expression, blocked cell cycle, inhibited human 20S proteasome	caspase3/4/5/10, 20S proteasome	(Czerwonka et al., 2020)
Earthworm extract	–	In vivo	Epinephrine induced myocardial infarction in rats	60 mg/kg/7 days	improved cardiac, liver and renal biomarkers such as creatine kinase (CK), albumin, creatinine, uric acid and repaired the antioxidant system	CK	(Oma et al., 2022)
Earthworm extract	Pheretima aspergillum	In vitro	LPS induced H9c2 cells apoptosis	62.5, 125, 250 μg/ml/1 hour	Increased the expression of Bcl2 and Bcl-x, and inhibited the expression of TNF-α, caspase3/8/9, t-Bid and Bax	Bcl2, Bcl-x, TNF-α, caspase3/8/9, t-Bid, Bax	(Li et al., 2015)
Dilong extract	Pheretima aspergillum	In vitro	High-KCl cardioplegic solution induced H9c2 cells apoptosis	31.25, 62.5, 125, 250 mg/ml/24 hours	Inhibitied the activation of IGF-I/IGF-IR/ERK pathway and the expression of uPA, Sp-1 and CTGF, inhibitied the expression of caspase-3 through MEK	uPA, Sp-1, CTGF, caspase-3, IGF-I/IGF-IR/ERK pathway, MEK pathway	(Han et al., 2014)
Earthworm extract	Pheretima aspergillum	In vitro	KCL induced H9c2 cells apoptosis	31.25, 62.5, 125, 250mg/ml/24 hours	Inhibited the expression of t-Bid, p-ERK1/2, uPA, SP1, CTGF, promoted the expression of Bcl2 and Bcl-x, and activated the IGF1R/PI3K/Akt signaling pathway	t-Bid, p-ERK1/2, uPA, SP1, CTGF, Bcl2, Bcl-x, IGF1R/PI3K/Akt signaling pathway	(Huang et al., 2019)
Lumbrokinase	Canada RNA biochemical Inc., Richmond, BC, Canada	in vivo	I-R injury in male Sprague-Dawley rats	10 μg/kg	Activated Sirt1, inhibited the protein expression of Bax and Cleaved Caspase-3, upregulated the protein expression of Bcl-2, alleviated myocardial I-R injury and decreased the induced expression of COX-2, iNOS and NF-κB	Sirt1, Bax, Cleaved Caspase-3, Bcl-2, COX-2, iNOS, NF-κB	(Wang et al., 2018)
Lumbrokinase	Harbin Jiuxin Science and Technology Industrial	in vivo	I-R injury in male Sprague-Dawley rats	1.2mg/kg, twice/week, a month	Inhibited ERK1/2 mediated activation of uPA/MMP and SP/CTGF fibrotic signaling pathways	ERK1/2, uPA/MMP and SP/CTGF signaling pathways	(Lai et al., 2015)
Dilong extract	Pheretima aspergillum	In vitro	RSC96 Schwann cells	125 μg/ml/24 hours	Mediated phosphorylation of PI3K/Akt pathway through the induction of IGF - I, stimulated the increase of PCNA, promoted cell proliferation	IGF-1, PI3K/Akt pathway	(Chang et al., 2011)
Earthworm extract	–	In vitro	PC12 cells	31.5, 125, 500 g/ml	Increased the level of GAP-43 and synapsin I, promoted the nerve growth factor mediated neurite outgrowth	GAP-43, synapsin I	(Chen et al., 2010)
Earthworm extract	Ohira ii earthworms	In vivo	Mice	100 ml (50%, 100%)/kg/7 days	Reduced the levels of NE, 5-HT and nitric oxide synthase (NOS) in serum and brain of mice and increased the pain threshold of mice	NE, 5-HT, NOS	(Luo et al., 2018)
Lumbrokinase	Boluoker, CRNA, Canada	in vivo	STZ induced diabetic rats	300, 600, 1,200 g/kg/28 days	Stimulated the secretion of IL-1, NGF, PDGF and TGF-β and the expression of CGRP and the recruitment of macrophages	IL-1, NGF, PDGF, TGF-β, CGRP	(Lee et al., 2015)
Earthworm extract	Eisenia foetida	In vivo	Ova induced asthmatic mice	50, 100, 200 mg/kg/56days	Regulated the balance of MMPs/TIMP-1, inhibited the infiltration of inflammatory cells in the lungs, and reduced the levels of Eot, IL-4, IL-5, IL-13 and plasma OVA-specific IgE	MMPs/TIMP-1, Eot, IL-4, IL-5, IL-13	(Zhang et al., 2021)
Earthworm protein	–	In vivo	STZ induced diabetic rats	0.045, 0.09, 0.18 g/kg/28 days	Inhibited oxidative stress, increased serum T level and activated NO receptor cGMP signal pathway	NO receptor cGMP signal pathway	(Zhang et al., 2023)
Earthworm protein	Baoji Fangsheng Biological Development Co., Ltd.	In vivo	STZ combined with high-fat diet induced diabetic rat.	45, 90, 180 mg/kg/28 days	Inhibited the transformation of CCSMC from “contractile” to “synthetic (proliferation)”	–	(Ji et al., 2024)
Earthworm protein	Baoji Fangsheng Biological Development Co., Ltd.	In vivo	STZ induced diabetic rats	45, 90, 180 mg/kg/28 days	Inhibited the expression of NF-κB, IL-1β and TNF-α, increased the expression of SOD and Nrf2	NF-κB, IL-1β, TNF-α, SOD, Nrf2	(Zhang et al., 2023)
Glycolipoprotein extract (G90)	Eisenia foetida	In vivo	Alloxan-induced diabetic rats	-/21 days	Promoted the formation of extracellular matrix, increased the fibroblast proliferation and neovascularization, collagen synthesis and early epithelial layer formation	–	(Goodarzi et al., 2016)
Glycoprotein extract (PvE-3)	Pheretima vulgaris	In vitro and vivo	NIH3T4 cell line and db/db mice, db/m mice	40, 80, 160, μg/ml/and 100μl(160mg/ml)/3, 7, 14 days	Promoted cell cycle progression from G0 phase to G1/S/G2/M and preserved ΔΨm	–	(Wang et al., 2023)

Table 1. Bioactivity of EE and mechanism

Sample

Source/Variety

In vitro/in vivo

Model

Dose/time of duration

mechanism

target/pathway

ref

Notes: “–” represents the source or dose was not shown. For lumbrokinase, we show the source instead of variety.

Lumbrokinase

Canada RNA biochemical Inc., Richmond, BC, Canada

In vivo

I-R injury in male Sprague-Dawley rats

1,10 μg/kg

Inhibited TLR4 expression, the phosphorylation of JNK, NF-κB, IκBα and the protein expression of COX-2, iNOS and MMP-9

TLR4, JNK, NF-Κb, IκBα, COX-2, iNOS, MMP9

(Wang et al., 2016)

Lumbrokinase

Canada RNA biochemical Inc., Richmond, BC, Canada

In vivo

mice

0.1, 1 mg/kg/7 days

Decreased the expression of IRE1 and inhibited the expression of transcription factors, XBP-1, caspase-12 and NF-κB, reduced NLRP3 inflammasome

IRE1, XBP-1, NF-κB, NLRP3, caspase-12

(Wang et al., 2022)

Earthworm active protein

Xian Guanmao Biotechnology Co., Ltd.

In vivo

Hyperlipidemic SD rats

50, 100, 200 mg/kg/28 days

Increased lipase activity and enhanced intestinal cholesterol metabolism transformation

–

(Yuan et al., 2018)

Polysaccharide-protein complex (PPC-DV)

Dendrobaena veneta

In vitro

Blood from peripheral vein

25, 50, 100 μg/ml

Targeted the P2Y12 receptor, COX-1 and PAR-1 pathways

P2Y12R, COX-1, PAR-1

(Poniedzialek et al., 2022)

Dilong extract

Pheretima vulgaris

In vivo and vitro

Collagen-induced arthritis in mice and murine RAW 264.7 macrophages

5, 10 g/kg/28 days and 75, 150, 300 μg/kg/1 days

Inhibited the NF-κB p65 activation, regulated the ratio of Th1/Th2 cells and improved the abnormal levels of inflammatory cytokines

NF-κB p65

(Bao et al., 2022)

Earthworm extract

El-Bagour, Menoufia, Egypt

In vivo

Acrylamide induced reproductive Injury in male rats

300 mg/ml/15 days

Inhibited the expression of P53 and Ki67, restored testicular antioxidant balance

P53, Ki67

(Ahmed et al., 2022)

Lumbrokinase

–

in vivo

I-R induced testicular injury in rats

80mg/kg

Inhibited the expression of Bax

Bax

(Danarto et al., 2019)

Earthworm extract

–

In vivo

Mouse wound model

0.1 ml

Reduced the ill-effects of inflammation, accelerated the secretion of hydroxyproline and TGF-β, increased the synthesis of collagen, and promoted the proliferation of blood capillary, and fibroblast. Accelerated the removal of dead tissue and foreign bodies by accelerating the production of IL- 6, white blood cells and platelets

TGF-β, IL- 6

(Deng et al., 2018)

Earthworm active ingredients

Eisenia foetida

In vitro

Tunicamycin induced L-02 cells apoptosis

0.1, 0.2, 0.4 mg/L/24 hours

Inhibited mRNA and protein expressions of GRP78, PERK, ATF4, Elf2α, CHOP, Bax, relieved ERS, he protein and mRNA expressions of Bcl-2, promoted the proliferation of L-02 cells.

GRP78, PERK, ATF4, Elf2α, CHOP, Bax, ERS, Bcl-2

(Duan et al., 2018)

Earthworm extract

Allolobophora caliginosa

In vivo

SiNPs induced liver injury in male albino rats

500, 1,000 mg/kg/30 days

Regulated the antioxidant balance of the liver

–

(Sadek et al., 2016)

Earthworm coelomic fluid

Allolobophora caliginosa

In vivo

Gentamicin induced hepatorenal toxicity in rats

45 mg/kg/7 days

Regulated the balance of oxidative system in the body

–

(Mohamed et al., 2020)

Earthworm extract

Perionyx excavates

In vivo

HFD-induced non-alcoholic fatty liver in guinea pigs

0.3, 1.4, 6.8 μg/kg

Inhibited the expression of TC, TG and LDL-C, alleviated liver injury in guinea pigs

TC, TG, LDL-C

(Deng et al., 2021)

Earthworm extract

–

In vivo

Silica-induced pulmonary fibrosis in mice

600 U/7, 14, 28 days

Activated Nrf2 pathway, inhibited oxidative stress, alleviated silica-induced apoptosis of lung epithelial cells and EMT

Nrf2 pathway, EMT pathway

(Yang et al., 2016)

Dilong extract

Guang dilong

In vivo

Bleomycin-induced pulmonary fibrosis mice

1.175, 2.35, 4.7g/kg/14 days

Combined with TGF-β1, inhibited the expression of α-SAM

α-SAM

(Wang et al., 2019)

Pheretima protein P2

Guang dilong

In vitro and vivo

TGF-β induced abnormal proliferation in MRC5 cells and bleomycin induced pulmonary fibrosis in mice

13.125μg/ml/2h and 5mg/kg/22days

Inhibited the protein expression of smad2/3 and p-smad2/3, inhibited the expression of α-SMA, Vimentin and Collagen I and increased the expression of E-cadherin

smad2/3, p-smad2/3, α-SMA, Vimentin, Collagen I, E-cadherin

(Li et al., 2022)

Water extract of earthworms

Pheretima aspergillum

In vivo and vitro

CCL4- induced liver fibrosis in mouse and TGFβ1 treated LX-2 HSCs and AML-12 cells

100,200 mg/kg/7 days and 30, 100 μg/ml/24 hours

Enhanced AMPK/GSK3β/Nrf2 cascade by activating LKB1, inhibited HSC activation, hepatocyte apoptosis and ferroptosis

LKB1, AMPK/GSK3β/Nrf2 cascade

(Zhang et al., 2024)

Earthworm granulation tissue extract

Eisenia fetida

In vivo

STZ induced diabetic rats

50, 70%/21 days

Scavenged free radicals, inhibited inflammatory pathways, reduced the levels of TNF-α, IL-1β, LPO, IL-6, promoted the synthesis of connective tissue framework protein and the expression of growth promoters

TNF-α, IL-1β, LPO, IL-6

(Elkhalifa et al., 2024)

Earthworm extract

–

In vivo

burn wound model on skin of mice

0.1 ml/3, 7, 11, 15 days

decreased edema, suppressed fibrosis, activated angiogenesis and epithelial regeneration, inhibited scar formation, and reduced the risk of infection.

–

(He et al., 2021)

Earthworm extract

Pheretima aspergillum

In vitro

Osteoblast-like MG-63 cells and RAW 264.7 macrophage cells

500 ng/ml – 12 mg/ml/2 days

increased osteoblast proliferation and differentiation, matrix calcium deposition and the expression levels of alkaline phosphatase (ALP), osteopontin (OPN) and osteocalcin (OCN), reduced the tartrate-resistant acid phosphatase (TRAP) activity of osteoclasts

ALP, OPN, OCN, TRAP

(Fu et al., 2014)

Earthworm active protein

–

In vitro

NIH3T4 cell line

37.5, 75, 150 mg/ml

Activated the MEK/ERK signaling pathway, promoted the cell cycle from G1 phase to S phase, and promoted cell proliferation

MEK/ERK signaling pathway

(Song et al., 2016)

Earthworm extract

–

In vitro

Human ovarian cancer cells A2780

400, 800 mg/ml/72 hours

Inhibited wnt / βcatenin pathway and promoted apoptosis

wnt / βcatenin pathway

(Chen et al., 2023)

Earthworm powder

Lumbricus terrestris

In vitro

Breast cancer cells (MCF-7) and prostate cancer cells (PC-3)

100, 200, 400 μg/ml/2 4 hours

Changed the permeability of the mitochondrial membrane and released cytochrome c

–

(Shafi and Faleh, 2019)

Earthworm extract

Perionyx excavates

In vivo

S180 tumor-bearing mice

30, 60, 90 mg/kg/25 days

Reduced the level of LDH, regulated the expression of Bax and Bcl2 protein

LDH, Bax, Bcl2

(Deng et al., 2019)

Earthworm coelomic fluid

Dendrobaena veneta

In vitro

A549 cells

125, 187.5, 250 μg/ml/24 hours

Increased expression of caspase3/4/5/10

caspase3/4/5/10

(Fiolka et al., 2019)

Protein-carbohydrate fraction (PE)

Dendrobaena veneta

In vitro

HT-29 cells

10–200 μg/ml

Inhibited mitochondrial metabolism, up-regulated caspase-3 expression, blocked cell cycle, inhibited human 20S proteasome

caspase3/4/5/10, 20S proteasome

(Czerwonka et al., 2020)

Earthworm extract

–

In vivo

Epinephrine induced myocardial infarction in rats

60 mg/kg/7 days

improved cardiac, liver and renal biomarkers such as creatine kinase (CK), albumin, creatinine, uric acid and repaired the antioxidant system

(Oma et al., 2022)

Earthworm extract

Pheretima aspergillum

In vitro

LPS induced H9c2 cells apoptosis

62.5, 125, 250 μg/ml/1 hour

Increased the expression of Bcl2 and Bcl-x, and inhibited the expression of TNF-α, caspase3/8/9, t-Bid and Bax

Bcl2, Bcl-x, TNF-α, caspase3/8/9, t-Bid, Bax

(Li et al., 2015)

Dilong extract

Pheretima aspergillum

In vitro

High-KCl cardioplegic solution induced H9c2 cells apoptosis

31.25, 62.5, 125, 250 mg/ml/24 hours

Inhibitied the activation of IGF-I/IGF-IR/ERK pathway and the expression of uPA, Sp-1 and CTGF, inhibitied the expression of caspase-3 through MEK

uPA, Sp-1, CTGF, caspase-3, IGF-I/IGF-IR/ERK pathway, MEK pathway

(Han et al., 2014)

Earthworm extract

Pheretima aspergillum

In vitro

KCL induced H9c2 cells apoptosis

31.25, 62.5, 125, 250mg/ml/24 hours

Inhibited the expression of t-Bid, p-ERK1/2, uPA, SP1, CTGF, promoted the expression of Bcl2 and Bcl-x, and activated the IGF1R/PI3K/Akt signaling pathway

t-Bid, p-ERK1/2, uPA, SP1, CTGF, Bcl2, Bcl-x, IGF1R/PI3K/Akt signaling pathway

(Huang et al., 2019)

Lumbrokinase

Canada RNA biochemical Inc., Richmond, BC, Canada

in vivo

I-R injury in male Sprague-Dawley rats

10 μg/kg

Activated Sirt1, inhibited the protein expression of Bax and Cleaved Caspase-3, upregulated the protein expression of Bcl-2, alleviated myocardial I-R injury and decreased the induced expression of COX-2, iNOS and NF-κB

Sirt1, Bax, Cleaved Caspase-3, Bcl-2, COX-2, iNOS, NF-κB

(Wang et al., 2018)

Lumbrokinase

Harbin Jiuxin Science and Technology Industrial

in vivo

I-R injury in male Sprague-Dawley rats

1.2mg/kg, twice/week, a month

Inhibited ERK1/2 mediated activation of uPA/MMP and SP/CTGF fibrotic signaling pathways

ERK1/2, uPA/MMP and SP/CTGF signaling pathways

(Lai et al., 2015)

Dilong extract

Pheretima aspergillum

In vitro

RSC96 Schwann cells

125 μg/ml/24 hours

Mediated phosphorylation of PI3K/Akt pathway through the induction of IGF - I, stimulated the increase of PCNA, promoted cell proliferation

IGF-1, PI3K/Akt pathway

(Chang et al., 2011)

Earthworm extract

–

In vitro

PC12 cells

31.5, 125, 500 g/ml

Increased the level of GAP-43 and synapsin I, promoted the nerve growth factor mediated neurite outgrowth

GAP-43, synapsin I

(Chen et al., 2010)

Earthworm extract

Ohira ii earthworms

In vivo

Mice

100 ml (50%, 100%)/kg/7 days

Reduced the levels of NE, 5-HT and nitric oxide synthase (NOS) in serum and brain of mice and increased the pain threshold of mice

NE, 5-HT, NOS

(Luo et al., 2018)

Lumbrokinase

Boluoker, CRNA, Canada

in vivo

STZ induced diabetic rats

300, 600, 1,200 g/kg/28 days

Stimulated the secretion of IL-1, NGF, PDGF and TGF-β and the expression of CGRP and the recruitment of macrophages

IL-1, NGF, PDGF, TGF-β, CGRP

(Lee et al., 2015)

Earthworm extract

Eisenia foetida

In vivo

Ova induced asthmatic mice

50, 100, 200 mg/kg/56days

Regulated the balance of MMPs/TIMP-1, inhibited the infiltration of inflammatory cells in the lungs, and reduced the levels of Eot, IL-4, IL-5, IL-13 and plasma OVA-specific IgE

MMPs/TIMP-1, Eot, IL-4, IL-5, IL-13

(Zhang et al., 2021)

Earthworm protein

–

In vivo

STZ induced diabetic rats

0.045, 0.09, 0.18 g/kg/28 days

Inhibited oxidative stress, increased serum T level and activated NO receptor cGMP signal pathway

NO receptor cGMP signal pathway

(Zhang et al., 2023)

Earthworm protein

Baoji Fangsheng Biological Development Co., Ltd.

In vivo

STZ combined with high-fat diet induced diabetic rat.

45, 90, 180 mg/kg/28 days

Inhibited the transformation of CCSMC from “contractile” to “synthetic (proliferation)”

–

(Ji et al., 2024)

Earthworm protein

Baoji Fangsheng Biological Development Co., Ltd.

In vivo

STZ induced diabetic rats

45, 90, 180 mg/kg/28 days

Inhibited the expression of NF-κB, IL-1β and TNF-α, increased the expression of SOD and Nrf2

NF-κB, IL-1β, TNF-α, SOD, Nrf2

(Zhang et al., 2023)

Glycolipoprotein extract (G90)

Eisenia foetida

In vivo

Alloxan-induced diabetic rats

-/21 days

Promoted the formation of extracellular matrix, increased the fibroblast proliferation and neovascularization, collagen synthesis and early epithelial layer formation

–

(Goodarzi et al., 2016)

Glycoprotein extract (PvE-3)

Pheretima vulgaris

In vitro and vivo

NIH3T4 cell line and db/db mice, db/m mice

40, 80, 160, μg/ml/and 100μl(160mg/ml)/3, 7, 14 days

Promoted cell cycle progression from G0 phase to G1/S/G2/M and preserved ΔΨm

–

(Wang et al., 2023)

**Table 2.** Earthworm peptides, their sequence, bioactivity and molecular mechanism
Peptide source	Peptide sequence	In vitro/in vivo	Model	Dose/time of duration	Molecular mechanism	target/pathway	ref
Notes:“–” represents no biological model was used, no dose or no molecular mechanism.
Earthworm protein autolysate	WNWLLPLMLG	In vitro	RAW 264.7 cells	500 μg/mL/24 hours	Targeted TLR2 and TLR4, promoted the secretion of cytokines	TLR2, TLR4	(Zhang et al., 2023)
Earthworm protein hydrolysate	AFWYGLPCKL, WPWQMSLY, GCFRYACGAAFY	In vitro	–	–	–	Angiotensin-converting enzyme	(He et al., 2021)
Earthworm protein in vitro gastrointestinal digestion product	SSPLWER and RFFGP	In vitro	–	–	–	Oxidative stress	(Lin et al., 2023)
Earthworm extracts	GRTGSTGATGPIGATGGSGLPGSNGATGIQGPMGRTGSTGPIGSTGATGAT	In vitro and vivo	NIH-3T3 cells and mice	0.45, 0.9, 1.8 μg/ml/12h and -/1, 3, 6, 10days	Activated the integrin α2β1 and RAS/MAPK signal pathways	integrin α2β1 and RAS/MAPK signal pathways	(Du et al., 2021)
Earthworm Lumbricus terrestris	NH 2-RNRRWCIDQQA	In vitro	SH-SY5Y cells	10 μg/ml/2, 4, 8 hours	Promoted ubiquitination degradation of p27 ^{Kip 1} protein	p27 ^{Kip 1}	(Kim et al., 2014)
Earthworm Lumbricus terrestris	NH 2-RNRRWCIDQQA	In vitro and in vivo	MNSCs and mouse	10 μg/ml/1 hour and 10 μg/20 days	Promoted ubiquitination degradation of p27 ^{Kip 1} protein	p27 ^{Kip 1}	(Kim et al., 2015)
Earthworm Lumbricus terrestris	QLICWRRFR-NH 2	In vitro and in vivo	BV-2 cells, SH-SY5Y cells and mice	20, 40, 60, 80 μg/ml/1 hour and 1, 5 mg/ml/3 days	Inhibited the activation of MAPK and AKT/NF-κB, reduced the levels of NO, iNOS, COX-2, TNF-α, IL-1β and IL-6	MAPK, AKT/NF-κB, NO, iNOS, COX-2, TNF-α, IL-1β, IL-6	(Seo et al., 2017)
Earthworm coelomic fluid	VQ-5 (VSSVQ) and AQ-5 (AMADQ)	In vivo	Mice	1.25, 2.5, 5 mg/kg	Reduced the levels of TNF-α and COX-2, inhibited the phosphorylation of IKKα/β, JNK and Erk (only AQ-5)	TNF-α, COX-2, IKKα/β, JNK, Erk	(Li et al., 2017)
Pheretima guillelmi skin	FSRYARMRDSRPWSDRKNNYSGPQFTYPPEKAPPEKLIKWNN EGSPIFEMPAEGGHIEP	In vitro	human and rabbit red blood cells	200 μg/ml	–	–	(Li et al., 2011)
Eisenia andrei	RIWWSGGWRRWRRW	In vitro	Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus)	5, 10, 20, 40, 80μg/ml	Killed E. coli through its secondary response, Killed S. aureus though membrane damage	–	(Wu et al., 2023)

Table 2. Earthworm peptides, their sequence, bioactivity and molecular mechanism

Peptide source

Peptide sequence

In vitro/in vivo

Model

Dose/time of duration

Molecular mechanism

target/pathway

ref

Notes:“–” represents no biological model was used, no dose or no molecular mechanism.

Earthworm protein autolysate

WNWLLPLMLG

In vitro

RAW 264.7 cells

500 μg/mL/24 hours

Targeted TLR2 and TLR4, promoted the secretion of cytokines

TLR2, TLR4

(Zhang et al., 2023)

Earthworm protein hydrolysate

AFWYGLPCKL, WPWQMSLY, GCFRYACGAAFY

In vitro

–

Angiotensin-converting enzyme

(He et al., 2021)

Earthworm protein in vitro gastrointestinal digestion product

SSPLWER and RFFGP

In vitro

–

Oxidative stress

(Lin et al., 2023)

Earthworm extracts

GRTGSTGATGPIGATGGSGLPGSNGATGIQGPMGRTGSTGPIGSTGATGAT

In vitro and vivo

NIH-3T3 cells and mice

0.45, 0.9, 1.8 μg/ml/12h and -/1, 3, 6, 10days

Activated the integrin α2β1 and RAS/MAPK signal pathways

integrin α2β1 and RAS/MAPK signal pathways

(Du et al., 2021)

Earthworm Lumbricus terrestris

NH 2-RNRRWCIDQQA

In vitro

SH-SY5Y cells

10 μg/ml/2, 4, 8 hours

Promoted ubiquitination degradation of p27 ^{Kip 1} protein

p27 ^{Kip 1}

(Kim et al., 2014)

Earthworm Lumbricus terrestris

NH 2-RNRRWCIDQQA

In vitro and in vivo

MNSCs and mouse

10 μg/ml/1 hour and 10 μg/20 days

Promoted ubiquitination degradation of p27 ^{Kip 1} protein

p27 ^{Kip 1}

(Kim et al., 2015)

Earthworm Lumbricus terrestris

QLICWRRFR-NH 2

In vitro and in vivo

BV-2 cells, SH-SY5Y cells and mice

20, 40, 60, 80 μg/ml/1 hour and 1, 5 mg/ml/3 days

Inhibited the activation of MAPK and AKT/NF-κB, reduced the levels of NO, iNOS, COX-2, TNF-α, IL-1β and IL-6

MAPK, AKT/NF-κB, NO, iNOS, COX-2, TNF-α, IL-1β, IL-6

(Seo et al., 2017)

Earthworm coelomic fluid

VQ-5 (VSSVQ) and AQ-5 (AMADQ)

In vivo

Mice

1.25, 2.5, 5 mg/kg

Reduced the levels of TNF-α and COX-2, inhibited the phosphorylation of IKKα/β, JNK and Erk (only AQ-5)

TNF-α, COX-2, IKKα/β, JNK, Erk

(Li et al., 2017)

Pheretima guillelmi skin

FSRYARMRDSRPWSDRKNNYSGPQFTYPPEKAPPEKLIKWNN EGSPIFEMPAEGGHIEP

In vitro

human and rabbit red blood cells

200 μg/ml

–

(Li et al., 2011)

Eisenia andrei

RIWWSGGWRRWRRW

In vitro

Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus)

5, 10, 20, 40, 80μg/ml

Killed E. coli through its secondary response, Killed S. aureus though membrane damage

–

(Wu et al., 2023)