Invest Clin 67(1): 19 - 29, 2026 https://doi.org/10.54817/IC.v67n1a02

Corresponding author. Zengguang Chen. Department of Cardiology, The Second People’s Hospital of Changzhou,

The Third Affiliated Hospital of Nanjing Medical University,No.68, Ge Hu Middle Road, Wujin District, China, Chang-

zhou, Jiangsu, 213000, China. Email:zengguangchen@126.com.

The impact of complement C1q/tumor

necrosis factor-related protein 6-mediated

cardiomyocyte pyroptosis on myocardial

fibrosis in rats with myocardial infarction.

Qiu Zhang, Xin Chen, Hao Pan and Zengguang Chen

Department of Cardiology, The Second People’s Hospital of Changzhou, The Third

Affiliated Hospital of Nanjing Medical University, Changzhou, JiangSu, China.

Keywords: Myocardial Infarction; Pyroptosis; Fibrosis; Inflammatory Response.

Abstract. Complement C1q/tumor necrosis factor-related protein 6

(CTRP6) has anti-inflammatory and metabolic regulatory properties, but its

role in ameliorating post-myocardial infarction (MI) myocardial fibrosis via

pyroptosis inhibition is unclear. This study investigated whether CTRP6 im-

proves post-MI myocardial fibrosis and cardiac dysfunction by suppressing car-

diomyocyte pyroptosis through the NLRP3/caspase-1/GSDMD pathway. Thirty

Sprague-Dawley rats were randomized to sham-operated (Sham), MI model

(MI), or CTRP6-treated (MI+CTRP6) groups. MI was induced by left anterior

descending coronary artery ligation; MI+CTRP6 rats received daily subcutane-

ous recombinant CTRP6 (0.2 mg/kg) from day 3 post-surgery for 28 days. Car-

diac function, fibrosis markers, pyroptosis-related proteins, and inflammatory

cytokines were assessed via Western blot, Masson staining, and ELISA. CTRP6

expression was lower in MI vs. Sham (p<0.05). CTRP6 treatment restored its

expression, reduced fibrosis markers and collagen deposition, and improved

cardiac function (p<0.05). It also downregulated pro-inflammatory cytokines

and increased anti-inflammatory cytokines (p<0.05). In other words, exoge-

nous CTRP6 ameliorated fibrosis and cardiac function by directly inhibiting the

NLRP3/caspase-1/GSDMD pyroptosis pathway.

20 Zhang et al.

Investigación Clínica 67(1): 2026

Impacto de la piroptosis de cardiomiocitos mediada

por CTRP6 en la fibrosis miocárdica en ratas con infarto

de miocardio.

Invest Clin 2026; 67 (1): 19 – 29

Palabras clave: Infarto de Miocardio; Piroptosis; fibrosis; Respuesta Inflamatoria.

Resumen. La proteína 6 relacionada con el factor de necrosis del comple-

mento C1q/tumor (CTRP6) tiene propiedades antiinflamatorias y reguladoras

metabólicas, pero su papel en la reducción de la fibrosis del miocardio postinfar-

to (MI) mediante la inhibición de la piroptosis no está claro. Este estudio inves-

tigó si el CTRP6 mejora la fibrosis miocárdica post-MI y la disfunción cardíaca

al suprimir la piroptosis de cardiomiocitos mediante la vía NLRP3/caspasa-1/

GSDMD. Treinta ratas Sprague-Dawley se asignaron aleatoriamente a grupos

operados con simulación (Sham), modelo MI (MI) o tratados con CTRP6 (MI +

CTRP6). El MI fue inducido por ligadura de la arteria coronaria descendente an-

terior izquierda. Las ratas MI+CTRP6 recibieron CTRP6 recombinante por vía

subcutánea diaria (0,2 mg/kg) a partir del día 3 tras la cirugía durante 28 días.

La función cardíaca (ecocardiografía), los marcadores de fibrosis, las proteínas

relacionadas con la piroptosis y las citocinas inflamatorias se evaluaron median-

te transferencia Western, tinción de Masson y ELISA. La expresión de CTRP6

fue menor en MI que en Sham (p<0,05). El tratamiento con CTRP6 restableció

su expresión, redujo los marcadores de fibrosis y de deposición de colágeno, y

mejoró la función cardíaca (p<0,05). También disminuyó la regulación de las

citocinas proinflamatorias y aumentó la regulación antiinflamatoria (p<0,05).

El CTRP6 protege contra la fibrosis miocárdica post-MI inhibiendo la piroptosis

de cardiomiocitos a través de la vía NLRP3/caspasa-1/GSDMD, reduciendo las

citocinas proinflamatorias y la activación de fibroblastos.

Received: 12-08-2025 Accepted: 20-10-2025

INTRODUCTION

Myocardial infarction (MI) represents

one of the most life-threatening cardiovas-

cular disorders globally, with its pathological

hallmark being the ischemic necrosis of car-

diomyocytes followed by progressive fibrosis,

which ultimately culminates in heart failure

and life-threatening arrhythmias 1. Epide-

miological data reveal a striking age-depen-

dent incidence pattern, with MI affecting ap-

proximately 3.8% of the population below 60

years compared to 9.5% in individuals over

60 years 2. While contemporary therapeutic

strategies, including revascularization pro-

cedures and pharmacological interventions,

have markedly enhanced acute-phase surviv-

al rates, the inexorable advancement of myo-

cardial fibrosis continues to pose significant

challenges to long-term patient outcomes

and quality of life 3. The scientific commu-

nity has recently focused on elucidating the

contributions of programmed cell death

pathways to myocardial injury pathophysiol-

ogy 4. Among these, pyroptosis—a lytic form

of cell death mediated by inflammasome

activation—exacerbates local inflammatory

responses and drives fibroblast activation

Effect of complement C1q/tumor necrosis factor-related protein 6 on myocardial infarction 21

Vol. 67(1): 19 - 29, 2026

through the release of pro-inflammatory cy-

tokines such as IL-1β and IL-18, potentially

serving as a key trigger for myocardial fibro-

sis 5. Despite these insights, the regulatory

network of pyroptosis in MI and its molec-

ular link with fibrosis remain incompletely

elucidated.

Within the complement C1q/tumor ne-

crosis factor-related protein (CTRP) family,

CTRP6 has gained prominence as a multi-

functional regulator, demonstrating simulta-

neous involvement in both glucolipid metab-

olism and inflammatory modulation, thereby

positioning it as a molecule of particular

interest in metabolic syndrome research 6.

Emerging evidence indicates that CTRP6 ex-

erts protective effects against vascular endo-

thelial inflammation by inhibiting the NF-κB

signaling cascade and exhibits anti-fibrotic

properties in the context of diabetic cardio-

myopathy 7. Notably, in myocardial ischemia-

reperfusion (I/R) injury models, CTRP6

expression levels correlate positively with

cardiomyocyte survival rates 8. However, di-

rect evidence on whether CTRP6 influences

myocardial fibrosis by modulating pyroptosis

remains lacking. The study by Liang et al.9

established that serum CTRP6 levels were

substantially diminished in I/R-injured rats

and inversely associated with both pyropto-

sis markers (GSDMD-N and caspase-1) and

the extent of collagen deposition, suggest-

ing a potential role for CTRP6 in restraining

fibrosis via pyroptosis inhibition. Neverthe-

less, the specific involvement of CTRP6 in MI

pathophysiology has not yet been systemati-

cally investigated.

In this study, we present a novel con-

ceptual framework that proposes a “CTRP6-

pyroptosis-fibrosis” regulatory axis in MI.

The primary objective is to delineate the

molecular mechanisms by which CTRP6

attenuates cardiomyocyte pyroptosis by

modulating the NLRP3/caspase-1/GSDMD

signaling pathway, thereby mitigating fibro-

blast activation and pathological collagen

accumulation. The findings are expected to

yield two significant contributions: first, the

identification of new therapeutic targets for

combating post-MI fibrotic complications;

and second, the establishment of a robust

theoretical platform that supports the trans-

lational potential of CTRP6, moving it from

its traditional recognition as a metabolic

modulator to its emerging role as a cardio-

vascular protective agent.

MATERIALS AND METHODS

Animal subjects

A total of thirty male Sprague-Dawley

(SD) rats, aged 4–6 weeks and weighing 200

± 20 g, were obtained from Beijing Vital-

star Biotechnology Co., Ltd. (License No.:

SCXK2023-0014). The animals were housed

under standardized conditions, with free ac-

cess to water, in a controlled environment

(temperature: 20–25°C; relative humidity:

60–70%) under a 12-hour light/dark cycle.

Following a one-week acclimatization period,

the experiments were conducted in compli-

ance with the 3Rs (Replacement, Reduction,

Refinement) principles and were approved

by the Institutional Animal Ethics Commit-

tee (No. 20210301002).

MI model establishment

The rats were randomly allocated into

three groups (n=10 per group). In two of

the groups, MI was surgically induced. Brief-

ly, the rats were anesthetized, secured in a

supine position, and intubated, after which

mechanical ventilation was initiated. After

thoracotomy, the heart was gently extruded,

and the left anterior descending coronary

artery was rapidly ligated approximately

2–3 mm distal to the aortic root, near the

junction between the left auricle and the

pulmonary artery cone, for MI modeling 10.

The MI model was considered successful if

the left ventricular ejection fraction (LVEF)

measured by Doppler echocardiography was

<50%. The third group underwent an iden-

tical surgical procedure without left ante-

rior descending ligation and served as the

sham group. Among the two MI groups, one

22 Zhang et al.

Investigación Clínica 67(1): 2026

received daily subcutaneous injections of

recombinant human CTRP6 (0.2 mg/kg)

starting three days post-MI induction and

continuing for 28 consecutive days (CTRP6

group). The other MI group received an

equivalent volume of saline and was desig-

nated the model group.

Evaluation of cardiac function

Upon completion of the treatment

regimen, transthoracic echocardiography

was performed on all animals using a high-

resolution color Doppler ultrasound system

equipped with a 15 MHz linear transducer.

Key cardiac functional parameters, includ-

ing LVEF, left ventricular fractional short-

ening (LVFS), left ventricular end-diastolic

diameter (LVEDD), and left ventricular end-

systolic diameter (LVESD), were recorded

for comparative analysis.

Western blot analysis of myocardial

protein expression

Following euthanasia, myocardial tissue

samples were immediately collected, and to-

tal protein was extracted by homogenizing

the tissues in a RIPA lysis buffer at a 1:5 ra-

tio, followed by incubation at 4°C for 40 min-

utes. Protein concentrations were measured

using the bicinchoninic acid (BCA) protein

assay kit. Then, equal amounts of protein

(30 μg per sample) were separated with 10%

SDS-PAGE and transferred to polyvinylidene

difluoride (PVDF) membranes using a wet

transfer system. After blocking for two hours

at room temperature, the membranes were

incubated overnight at 4°C with primary

antibodies (all diluted 1:1000 in blocking

buffer): anti-CTRP6 (ab300583, Abcam),

anti-Collagen III (ab7535, Abcam), anti-α-

smooth muscle actin (α-SMA) (ab314895,

Abcam), anti-transforming growth factor-β1

(TGF-β1) (ab315254, Abcam), anti-NOD-

like receptor family pyrin domain containing

3 (NLRP3) (ab263899, Abcam), anti-cl-Cas-

pase-1 (ab198447, Abcam), anti-gasdermin

D (GSDMD) (ab219800, Abcam), anti-N

terminal (NT)-GSDMD (ab215203, Abcam),

anti-interleukin (IL)-1β (ab315084, Abcam),

and GAPDH (ab8245, Abcam). Subsequently,

the membranes were incubated with appro-

priate horseradish peroxidase (HRP)-conju-

gated secondary antibodies (goat anti-rabbit

or goat anti-mouse IgG, 1:5000 dilution,

ab308009, Abcam) for two hours at room

temperature. Using enhanced chemilumi-

nescence (ECL) detection reagents, protein

bands were visualized and photographed. Fi-

nally, relative protein expression levels were

quantified.

Histopathological examination

of myocardial tissues

Cardiac tissues were fixed in 10% para-

formaldehyde for 24 hours, then processed

through a graded ethanol series for dehy-

dration. After xylene clearing, tissues were

embedded in paraffin and sectioned at 3-5

μm thickness. Subsequently, sections were

stained with hematoxylin and eosin (H&E)

and examined under a light microscope at

200× magnification to evaluate myocardial

morphology (focusing on muscle fiber ar-

rangement and ventricular wall thickness).

For myocardial fibrosis assessment, tissue

fixation and section preparation followed

the same procedures as for H&E staining.

The sections were then stained using a Mas-

son’s trichrome kit according to the manu-

facturer’s instructions. Fibrotic changes in

myocardial tissue were evaluated under light

microscopy at 200× magnification, with col-

lagen fibers appearing blue in the stained

sections.

Measurement of serum inflammatory

cytokines

Blood samples (3 mL) were obtained

from the abdominal aorta into sterile tubes,

and serum was separated by centrifugation.

Serum levels of IL-1β (CSB-E08055r-IS),

tumor necrosis factor-alpha (TNF-α) (CSB-

E11987r), IL-6 (CSB-E04640r), IL-8 (CSB-

E07451r), and IL-10 (CSB-E04595r) were

measured using commercially available en-

zyme-linked immunosorbent assay (ELISA)

Effect of complement C1q/tumor necrosis factor-related protein 6 on myocardial infarction 23

Vol. 67(1): 19 - 29, 2026

kits (Wuhan Huamei Biological Engineering

Co., LTD.) according to the manufacturer’s

instructions.

Endpoints

The endpoints of this study included (1)

a quantitative assessment of CTRP6 protein

expression levels in myocardial tissues after

MI, and (2) a comprehensive evaluation of

CTRP6’s therapeutic effects on myocardial

fibrosis, histopathological damage, cardiac

function, and pyroptosis.

Statistical analysis

All statistical analyses were performed

using SPSS version 25.0 (IBM Corp., USA).

Continuous variables with normal distribu-

tion were presented as mean ± standard

deviation ( ± sd). An independent-samples

t-test was used to compare the two groups.

For multiple-group comparisons, one-way re-

peated-measures analysis of variance (ANO-

VA) was employed, followed by Fisher’s least

significant difference (LSD) post hoc test for

pairwise comparisons. A two-tailed p of less

than 0.05 was considered statistically signifi-

cant for all analyses.

RESULTS

MI model establishment and CTRP6

expression

All rats in the sham group survived.

In both the CTRP6 and model groups, one

rat died in each group, while the surviving

rats successfully met the MI model criteria

(LVEF <50%). Western blot analysis showed

that CTRP6 protein expression in myocardi-

al tissue was significantly lower in the model

group than in the sham group (p<0.001),

indicating downregulation of CTRP6 in MI

(Fig. 1).

CTRP6 attenuates myocardial fibrosis and

pathological damage in MI rats

Protein expression analysis showed that

although CTRP6 levels in the CTRP6 group

remained lower than those in the sham con-

trols, they were significantly higher than in

the model group (p<0.001), confirming suc-

cessful CTRP6 upregulation through recom-

binant human CTRP6 administration. Addi-

tionally, the model group displayed markedly

increased expression of fibrotic markers, in-

cluding Collagen III, α-SMA, and TGF-β1,

compared with the sham group (p<0.01).

Importantly, CTRP6 treatment significantly

reduced the expression of these fibrotic pro-

teins compared with the model group (p<

0.01), suggesting its potential to mitigate

the progression of myocardial fibrosis. Histo-

pathological examination with H&E staining

showed well-organized myocardial fibers and

intact tissue structure in the sham controls.

In contrast, the model group exhibited char-

acteristic pathological changes, such as dis-

organized myocardial fibers and ventricular

wall thinning. CTRP6 treatment markedly al-

leviated these abnormalities, resulting in in-

creased ventricular wall thickness and more

Fig. 1. CTRP6 protein expression in MI. An independent-samples t-test was used to compare groups (***p<0.001).

CTRP6:complement C1q/tumor necrosis factor-related protein 6. MI: Myocardial infarction.

Expression of CTRP6 protein

Sham group Model group

0.8

0.6

0.4

0.2

0.0

24 Zhang et al.

Investigación Clínica 67(1): 2026

organized fiber alignment compared with un-

treated MI rats. Masson’s trichrome staining

supported these results, with the sham group

showing minimal collagen deposition (indi-

cated by blue staining). The model group ex-

hibited extensive myocardial fibrosis, whereas

CTRP6-treated animals showed significantly

reduced collagen accumulation and better-

preserved myocardial structure (Fig. 2).

CTRP6 ameliorates cardiac dysfunction

in MI rats

The echocardiographic assessment re-

vealed significant cardiac dysfunction in the

MI model group compared to sham-operat-

ed controls. Specifically, the model group

exhibited markedly reduced LVEF and

LVFS, accompanied by increased LVEDD

and LVESD (p<0.05). Notably, CTRP6 ad-

ministration effectively attenuated these

pathological changes, with treated animals

demonstrating significantly improved LVEF

and LVFS, along with reduced LVEDD and

LVESD, compared with untreated MI rats

(p<0.05; Fig. 3).

CTRP6 attenuates pyroptotic cell death

in infarcted myocardium

Analysis of inflammatory mediators

showed a strong pro-inflammatory state in the

myocardial tissue of the model group, with sig-

nificantly increased levels of IL-1β, TNF-α, IL-6,

Fig. 2. Effect of CTRP6 on myocardial fibrosis and tissue damage in MI rats. (a) Measurement of myocardial

fibrosis-related protein expression in MI rats treated with exogenous CTRP6. (b) Hematoxylin and

eosin (H&E) staining of myocardial tissue (200×). (c) Masson staining of myocardial tissue (200×).

Repeated-measures analysis of variance and LSD intra-group tests were used to compare multiple

groups: ***p<0.001 compared with the sham group; **p<0.01 compared with the model group;

##p<0.01. Complement C1q/tumor necrosis factor-related protein 6 (CTRP6), α-smooth muscle

actin (α-SMA), and transforming growth factor-β1 (TGF-β1). The portion of inflammatory cell infil-

tration is highlighted in the figure. MI: Myocardial infarction.

Sham group (n=10)

Model group (n=9)

CTRP6 group (n=9)

Expression of CTRP6 protein

Expression of fibrotic protein

Sham group Model group CTRP6 group

0.8

0.6

0.4

0.2

0.0

1.0

0.4

0.3

0.2

0.1

0.0

0.5

Sham group Model group CTRP6 group

n=10 n=9 n=9

Collagen III α-SMA TGFβ-1

Sham group Model group CTRP6 group

Effect of complement C1q/tumor necrosis factor-related protein 6 on myocardial infarction 25

Vol. 67(1): 19 - 29, 2026

and IL-8, alongside decreased IL-10, compared

to sham controls (p<0.05). CTRP6 treatment

effectively adjusted this inflammatory imbal-

ance, significantly reducing pro-inflammatory

cytokines (IL-1β, TNF-α, IL-6, and IL-8) while

boosting anti-inflammatory IL-10 expression

(p<0.05). Regarding pyroptosis-related pro-

teins, western blot analysis revealed substantial

upregulation of NLRP3, cleaved Caspase-1, GS-

DMD, NT-GSDMD, and IL-1β in both the mod-

el and CTRP6 groups compared to the sham

group (p<0.05); however, their levels were

lower in the CTRP6 group than in the control

group (p<0.05). These findings suggest that

CTRP6 suppresses myocardial pyroptosis in MI

rats (Fig 4).

Fig. 3. Effect of CTRP6 on cardiac function (LVEF, LVFS, LVEDD, LVESD) in MI rats. Repeated-measures

analysis of variance and LSD intra-group tests were used to compare multiple groups; ***p<0.001.

Left ventricular ejection fraction (LVEF), left ventricular fractional shortening (LVFS), left ventricu-

lar end-diastolic diameter (LVEDD), and left ventricular end-systolic diameter (LVESD). MI: Myocar-

dial infarction; CTRP6: Complement C1q/tumor necrosis factor-related protein 6.

Fig. 4. Effect of CTRP6 on myocardial pyroptosis in MI rats. (a) Effect of CTRP6 on the levels of inflammatory factors

(IL-1β, TNF-α, IL-6, IL-8, IL-10) in myocardial tissues of MI rats. (b) Effect of CTRP6 on the expression of focal

death proteins (NLRP3, cl-Caspase-1, GSDMD, NT-GSDMD, IL-1β) in myocardial tissues of MI rats. Repeated-

measures analysis of variance and LSD intra-group tests were used to compare multiple groups, compared with

the sham group **p<0.01, and the model group ##p<0.01. NOD-like receptor family pyrin domain containing

3 (NLRP3), gasdermin D (GSDMD), N-terminal (NT), interleukin (IL), tumor necrosis factor-α (TNF-α). MI: Myo-

cardial infarction; CTRP6: Complement C1q/tumor necrosis factor-related protein 6.

Expression of fibrotic protein

Sham group (n=10)

Model group (n=9)

CTRP6 group (n=9)

Sham group (n=10)

Model group (n=9)

CTRP6 group (n=9)

Levels of inflammatory factors

(pg/mL)

Sham group Model group CTRP6 group

n=10 n=9 n=9

Sham group CTRP6 group

n=10 n=9 n=9

Model group

Sham group Model group CTRP6 group

n=10 n=9 n=9

Sham group CTRP6 group

n=10 n=9 n=9

Model group

LVEF (%)

LVEDD (mm)

LVFS (%)

LVESD (mm)

0.8

0.6

0.0

0.2

0.4

26 Zhang et al.

Investigación Clínica 67(1): 2026

DISCUSSION

Through MI modeling in rats and ex-

ogenous CTRP6 intervention experiments,

this study demonstrates that CTRP6 exerts

cardioprotective effects in MI via a novel

mechanism that suppresses cardiomyocyte

pyroptosis, thereby ameliorating myocardial

fibrosis and restoring cardiac function. These

findings not only substantiate the previously

hypothesized protective role of CTRP6 in

ischemic myocardial injury but also offer

novel therapeutic avenues by identifying the

pyroptosis-fibrosis axis as a potential target

for myocardial repair strategies. Specifically,

the therapeutic benefits of CTRP6 were: (1)

recovery of cardiac function: LVEF was im-

proved by subcutaneous injection of 0.2 mg/

kg/ day (28 days) in MI rats; (2) Anti-fibro-

sis: reduced collagen deposition and blocked

the TGF-β1 pathway.

In the present study, we initially ob-

served significantly decreased CTRP6 pro-

tein levels in the myocardial tissue of MI

model rats (Fig. 1), suggesting CTRP6’s

potential role in MI development and pro-

gression. This finding supports the find-

ings of Tabatabaei SA et al., who reported a

similar reduction in CTRP6 in patients with

coronary artery disease 11. The subsequent

administration of exogenous CTRP6 effec-

tively restored CTRP6 levels and yielded two

clinically relevant outcomes: a significant

reduction in fibrotic markers (Collagen

III, α-SMA, and TGF-β1) and improved car-

diac function parameters (LVEF and LVFS)

(Figs. 2-3). These linked effects provide pre-

liminary evidence for CTRP6’s therapeutic

potential in the treatment of MI. By analyz-

ing our data systematically and integrating

it with existing literature, we identified two

complementary mechanisms through which

CTRP6 likely exerts its cardioprotective ef-

fects: (1) Modulation of inflammation lead-

ing to reduced fibrosis: Our results clearly

show that CTRP6 treatment causes a signifi-

cant change in cytokine profiles, character-

ized by decreased levels of pro-inflammatory

mediators (IL-1β, TNF-α, IL-6, and IL-8) and

increased levels of the anti-inflammatory cy-

tokine IL-10 (Fig. 4). These findings extend

previous research 12 suggesting that CTRP6

may regulate inflammatory responses by

inhibiting the NF-κB signaling pathway, ul-

timately preventing NLRP3 inflammasome

activation. This mechanism would decrease

pro-inflammatory cytokine release and re-

duce inflammation in the myocardial en-

vironment. (2) Direct inhibition of fibrotic

pathways: Our study further confirms the

work of Yan et al. 13, showing that CTRP6

suppresses the TGF-β1/Smad3 pathway, pre-

venting the transformation of fibroblasts

into myofibroblasts. This is supported by our

observed reductions in TGF-β1 and α-SMA

protein levels, as well as notably less collagen

deposition (particularly Collagen III) in rats

treated with CTRP6. Notably, our findings

highlight the connection between CTRP6’s

anti-fibrotic effects and its ability to inhibit

pyroptosis 14. This connection is evident be-

cause IL-1β and IL-18, produced during py-

roptosis, are potent activators of fibroblasts

and promote excessive extracellular matrix

(ECM) production 15. Therefore, we propose

that CTRP6 coordinates a multi-faceted an-

ti-fibrotic approach by sequentially blocking

“pyroptosis → cytokine release → fibroblast

activation”. In addition, our study presents

the first experimental evidence that CTRP6

suppresses cardiomyocyte pyroptosis by

modulating the NLRP3/caspase-1/GSDMD

signaling pathway. The underlying mecha-

nisms may involve the following aspects:

Upstream regulation: CTRP6 likely initiates

its protective effects through activation of

the AMPKα pathway, which subsequently

attenuates reactive oxygen species (ROS)-

mediated NLRP3 inflammasome assembly 16.

Diminished ROS production reduces NLRP3

oligomerization, suppresses caspase-1 self-

cleavage (as evidenced by decreased cl-cas-

pase-1 expression), and ultimately prevents

GSDMD proteolytic activation (manifested

as reduced NT-GSDMD fragments) (Fig. 4).

Downstream effects: By inhibiting the pyrop-

Effect of complement C1q/tumor necrosis factor-related protein 6 on myocardial infarction 27

Vol. 67(1): 19 - 29, 2026

tosis executioner protein GSDMD, CTRP6

diminishes plasma membrane pore forma-

tion, thereby preventing the release of IL-1β

and IL-18 and consequently disrupting the

vicious cycle of “pyroptosis-inflammation-

fibrosis” (Fig. 4). While the current litera-

ture offers limited precedents for CTRP6’s

role in pyroptosis regulation, our findings

align with established clinical evidence that

identifies lipid peroxidation and membrane

disruption as fundamental prerequisites for

pyroptotic cell death 17,18. Given that CTRP6

functions as a lipid metabolism regulator, it

may enhance cellular resistance to pyropto-

sis by maintaining membrane phospholipid

homeostasis, such as by increasing sphin-

gomyelin levels 19. This intriguing hypoth-

esis warrants further investigation through

comprehensive lipidomic profiling in future

studies.

Furthermore, this study is the first to

establish the “CTRP6-pyroptosis-fibrosis”

regulatory axis, elucidating the molecu-

lar mechanism by which CTRP6 attenuates

post-MI fibrosis via targeting the NLRP3/

caspase-1/GSDMD pathway. Using a well-

established MI rat model, we demonstrated

that intravenous administration of exog-

enous CTRP6 improves cardiac function.

These findings provide crucial experimen-

tal evidence supporting the development

of CTRP6-based therapeutic approaches,

including both gene therapy and protein re-

placement strategies. Our research suggests

several promising clinical applications that

warrant further investigation: First, regular

monitoring of serum CTRP6 levels could po-

tentially serve as a valuable biomarker for

evaluating the risk of fibrosis development

in MI patients. Second, combinatorial regi-

mens pairing CTRP6 with established pyrop-

tosis inhibitors (such as MCC950) may pro-

duce enhanced anti-fibrotic effects through

synergistic mechanisms. Third, the dual pro-

tective properties of CTRP6—targeting both

metabolic and cardiovascular systems—may

be particularly beneficial for the manage-

ment of diabetic patients with MI, potential-

ly yielding superior therapeutic outcomes in

this high-risk population.

While our study provides important

insights, several limitations must be con-

sidered. First, the precise molecular mecha-

nism remains unclear—we cannot determine

whether CTRP6 exerts its anti-pyroptotic ef-

fects through direct interaction with NLRP3

or via indirect modulation of upstream regu-

latory kinases, such as ASK1. Second, our

experimental design did not incorporate

sex-based analyses, despite existing evidence

suggesting estrogen may influence CTRP6

expression patterns 20. Third, the absence of

clinical specimen data prevents validation of

the observed relationship between CTRP6

levels and pyroptosis markers.

As a conclusion, CTRP6 ameliorates myo-

cardial fibrosis and enhances cardiac function

by inhibiting NLRP3/caspase-1/GSDMD-me-

diated cardiomyocyte pyroptosis, thereby re-

ducing pro-inflammatory cytokine release and

fibroblast activation in an MI animal model.

These findings not only expand our understand-

ing of CTRP6’s role in cardiovascular diseases

but also provide novel strategies for precision

medicine in the treatment of MI.

Acknowledgements

Not applicable.

Funding

The authors declare that no funds,

grants, or other support were received du-

ring the preparation of this manuscript.

Ethical approval

The study protocol was approved by the

Ethics Committee of The Second People’s

Hospital of Changzhou (Approval num-

ber:20210301002).

Conflict of interest

The authors had no separate personal,

financial, commercial, or academic conflicts

of interest.

28 Zhang et al.

Investigación Clínica 67(1): 2026

Availability of data and material

All data generated or analyzed during

this study are included in this published ar-

ticle.

ORCID numbers of authors

• Qiu Zhang (QZ):

0009-0002-5874-7484

• Xin Chen (XC):

0009-0005-6572-9323

• Hao Pan (HP):

0009-0005-4335-9325

• Zengguang Chen (ZGC):

0000-0002-0895-3215

Author contributions

Study conception and design: ZGC, QZ;

Data collection: XC; Data analysis and inter-

pretation: HP; Drafting of the article: QZ.

Critical revision of the article: All authors.

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