Excessive mitochondrial fission in myocardial ischemia/reperfusion injury: Pathophysiological relevance and potential cardioprotective modulation by plant-derived interventions and electroacupuncture

Table 2 Experimental evidence for plant-derived interventions and electroacupuncture associated with mitochondrial fusion-fission-related changes in myocardial ischemia/reperfusion injury and related experimental models

Intervention type	Intervention	Composition or acupoints	Experimental model	Study groups	Sample size and design	Dosage, administration route, and regimen	Main findings	Evidence strength	Ref.
Defined combination	GRS	Ginsenoside Rb1 + ruscogenin + schisandrin (ratio: 6:0.75:6)	In vivo: ICR mice LAD I/R (30 minutes/24 hours). In vitro: H9c2 H/R (6 hours/6 hours; 2-hour reoxygenation for Drp1 and 1-hour for AMPK assays); NRVMs H/R (12 hours/12 hours)	In vivo: Sham; I/R; GRS + I/R; positive control (captopril). In vitro: Ctrl; H/R; GRS + H/R	In vivo: n = 6 (cardiac); R: Yes; B: Surgery	In vivo: 6.4 mg/kg, 12.8 mg/kg, 19.2 mg/kg; MED NR; administered i.p. within 10 minutes after reperfusion onset. In vitro: 0.1 μg/mL, 1 μg/mL, 10 μg/mL; MEC NR; added at hypoxia onset and maintained throughout H/R	In vivo: ↑: EF; ↓: Infarct size. In vivo/in vitro: ↑: Bcl-2/Bax ratio; ↓: P-Drp1(Ser616), LDH, caspase-3. In vivo/in vitro: ↑: P-AMPK	A, C; AMPK siRNA (C); Early Rep (A): Potential acute relevance; Isch (C)	Li et al[80]
PBM (injection)	Shenmai Injection	Aqueous extracts from Panax ginseng C.A. Mey (Araliaceae; Ginseng Radix et Rhizoma) and Ophiopogon japonicus (Thunb) Ker Gawl (Asparagaceae; Ophiopogonis Radix)	In vivo: SD rat MI model (LAD ligation). In vitro: H9c2 H/R (12 hours/2 hours)	In vivo: Sham; MI; MI + SMI; positive control (valsartan). In vitro: Ctrl; H/R; SMI + H/R	In vivo: n = 8 (EF/FS), 3 (infarct); R: Yes; B: Echocardiography	In vivo: 0.38 mL/kg; MED NR; post-MI treatment; SMI route/frequency NR (28-day protocol). In vitro: 1 μL/mL, 2.5 μL/mL, 5 μL/mL; MEC NR; pretreated for 24 hours before H/R	In vivo: ↑: EF, FS; ↓: Infarct size. In vitro: ↑: MFN1/2 and OPA1 mRNA, Parkin, Beclin 1, LC3-II/LC3-I, ΔΨm; ↓: Drp1 and Fis1 mRNA, mitochondrial fragmentation, cytosolic Ca2+ overload, mPTP opening	C, D; Assoc; Post-MI (D): Recovery; Pre (C)	Yu et al[74]
PBM (injection)	Guanxinning Injection	Extracts of Salvia miltiorrhiza Bunge (Lamiaceae; Salviae Miltiorrhizae Radix et Rhizoma) and Ligusticum chuanxiong Hort (Apiaceae; Chuanxiong Rhizoma)	Wistar rat LAD I/R (30 minutes/60 minutes) in a CRF background	Sham; CRF; I/R; CRF + I/R + IPost; CRF + I/R + IPost + GXN; CRF + I/R	n = 15 (allocated); R: Yes; B: NR	Administered i.v. 10 minutes after ischemia onset; dose NR; MED NR	AN/AAR: 0.17 ± 0.09 (GXN + IPost), 0.39 ± 0.07 (CRF + I/R), 0.29 ± 0.07 (IPost); mean ± SD. GXN + IPost vs CRF + I/R: ↑: MFN1, OPA1; ↓: Drp1, cTnT, CK-MB. GXN + IPost vs IPost alone: ↑: MFN1, OPA1; ↓: CK-MB	A; Assoc; Isch + IPost: Potential acute relevance (combined)	Wang et al[75]
PBM (capsule)	Shuangshen Ningxin Formula	Extracts of Panax ginseng C.A. Mey (Araliaceae; Ginseng Radix et Rhizoma) + Salvia miltiorrhiza Bunge (Lamiaceae; Salviae Miltiorrhizae Radix et Rhizoma) + Corydalis yanhusuo W.T.Wang ex Z.Y.Su & C.Y.Wu (Papaveraceae; Corydalis Rhizoma)	In vivo: SD rat LAD I/R (45 minutes/24 hours). In vitro: CMECs OGD/R (4 hours/2 hours)	In vivo: Sham; I/R; SSNX + I/R. In vitro: Ctrl; OGD/R; SSNX + OGD/R	In vivo: n = 6 (EF); cells: n = 6 (viability); R: Yes (rats); B: NR	In vivo: 45 mg/kg/day, 90 mg/kg/day, 180 mg/kg/day; MED NR; gavage for 7 days before I/R. In vitro: 25 μg/mL; MEC NR; added before OGD; lead time NR	In vivo: ↑: EF. In vivo/in vitro: ↓: NR4A1, Mff, Drp1, HK II, Cyt c, caspase-9	A, C; NR4A1 inhibitor comparator (C); Pre: Post-onset untested	Liu et al[83]
Botanical decoction	Taohong Siwu Decoction	Prunus persica (L) Batsch (Rosaceae; Persicae Semen) + Carthamus tinctorius L (Asteraceae; Carthami Flos) + Angelica sinensis (Oliv) Diels (Apiaceae; Angelicae Sinensis Radix) + Paeonia lactiflora Pall (Paeoniaceae; Paeoniae Radix Alba) + Ligusticum chuanxiong Hort (Apiaceae; Chuanxiong Rhizoma) + Rehmannia glutinosa (Gaertn) Libosch ex DC (Orobanchaceae; Rehmanniae Radix Praeparata)	SD rat MI (LAD ligation)	MI; THSWD + MI	n = 13 (allocated); R: Yes; B: Echocardiography	1 mL/day; MED NR; gavage for 4 weeks, starting 2 days after MI (reported concentration 1.13 g/mL)	4-week: ↓ LVESV; EF/FS differences NS. ↑: P-Akt/Akt. ↓: Fis1, mitochondrial ROS; LC3B: No statistically significant change	D; Assoc; Post-MI: Recovery	Luo et al[82]
PBM (pill)	Qishen Yiqi Dripping Pill	Astragalus membranaceus (Fisch) Bunge (Fabaceae; Astragali Radix) + Salvia miltiorrhiza Bunge (Lamiaceae; Salviae Miltiorrhizae Radix et Rhizoma) + Dalbergia odorifera T.C.Chen (Fabaceae; Dalbergiae Odoriferae Lignum) + Panax notoginseng (Burkill) F.H. Chen (Araliaceae; Notoginseng Radix et Rhizoma)	ICR male mice MI (LAD ligation)	Sham; MI; QSYQ + MI; positive control (enalapril)	n = 6 (LVEF); R: Yes; B: H&E assessment	12 mg/kg/day, 35 mg/kg/day, 105 mg/kg/day; MED NR; gavage for 14 days after MI	↑: LVEF, MFN2, L-OPA1/S-OPA1, PGC-1α, TFAM, NRF1. ↓: P-Drp1(Ser616)/Drp1	D; Assoc; Post-MI: Recovery	Wu et al[79]
PBM (capsule)	Danqi Soft Capsule	50% Panax notoginseng (Burkill) F.H. Chen (Araliaceae; Notoginseng Radix et Rhizoma) + 50% Salvia miltiorrhiza Bunge (Lamiaceae; Salviae Miltiorrhizae Radix et Rhizoma)	In vivo: SD rat LAD I/R (30 minutes/24 hours). In vitro: H9c2 H2O2 injury	In vivo: Sham; I/R; DQ + I/R. In vitro: Ctrl; H2O2; DQ + H2O2	In vivo: n = 4 (infarct), 6 (serum); R: Yes; B: Histology scoring	In vivo: 0.6 g/kg/day, 1.2 g/kg/day; MED NR; gavage for 5 days before I/R. In vitro: 300 mg/mL, 600 mg/mL; MEC NR; pretreated for 2.5 hours before H2O2 exposure	In vivo: ↑: Bcl-2; ↓: Infarct size, p-CaMKII, p-Drp1(Ser616), cTnT, LDH, cleaved caspase-3, Bax. In vitro: ↑: Bcl-2, ΔΨm; ↓: P-CaMKII, p-Drp1(Ser616), ROS, Cyt c, cleaved caspase-3, caspase-3/8/9 activities, Bax, mPTP opening	A, E; Assoc; Pre: Post-onset untested	Yang et al[76]
PBM (pill)	Tongmai Yangxin Pill	Multiple constituents, including Rehmannia glutinosa (Gaertn.) Libosch ex DC (Orobanchaceae; Rehmanniae Radix) + Spatholobus suberectus Dunn (Fabaceae; Spatholobi Caulis) + Ophiopogon japonicus (Thunb) Ker Gawl (Asparagaceae; Ophiopogonis Radix) + Glycyrrhiza uralensis Fisch ex DC (Fabaceae; Glycyrrhizae Radix et Rhizoma) + Reynoutria multiflora (Thunb) Moldenke (Polygonaceae; Polygoni Multiflori Radix Praeparata) + Schisandra chinensis (Turcz) Baill (Schisandraceae; Schisandrae Chinensis Fructus) + Codonopsis pilosula (Franch) Nannf (Campanulaceae; Codonopsis Radix) + Ziziphus jujuba Mill (Rhamnaceae; Jujubae Fructus) + Cinnamomum cassia (L) J.Presl (Lauraceae; Cinnamomi Ramulus) + Asini Corii Colla + Mauremydis Carapax et Plastrum	In vivo: SD rat LAD I/R (30 minutes/120 minutes). In vitro: H9c2 H/R (6 hours/24 hours)	In vivo: Sham; I/R; TMYX + I/R (1/2/4 g/kg); positive control (raloxifene). In vitro: Ctrl; H/R; TMYX + H/R	In vivo: n = 9 (EF), 3 (infarct), 8 (serum); R: Yes; B: NR	In vivo: 1 g/kg/day, 2 g/kg/day, 4 g/kg/day; MED NR; gavage for 7 days after I/R establishment per Methods; pretreatment wording elsewhere is inconsistent. In vitro: 100 μg/mL; MEC NR; timing NR	In vivo: ↑: EF, MFN1/2; ↓: Drp1, Fis1, ROS, CK-MB, cTnI, LDH. In vitro: ↑: ERα, PGC-1α, ATP, ΔΨm; ↓: ROS, Bax and caspase-3 mRNA	A, C; ERα siRNA (C); timing inconsistent (A): Relevance unclear; NR (C)	Yu et al[84]
Electroacupuncture	Electroacupuncture pretreatment	Acupoints: PC6 + ST36 + CV4	SD rat LAD I/R (30 minutes/120 minutes)	Sham; I/R; EA + I/R; EA + compound C + I/R; EA + ML385 + I/R	n = 3 (ROS/SOD/MDA), 6 (protein); R: Yes; B: NR	EA: 20 minutes/day for 7 days before I/R (2 Hz, 1 mA); inhibitors administered i.p. 30 minutes before model induction	Cardiac function and infarct size were not assessed. ↑: SOD, AMPK, NRF2. ↓: Drp1, Mff, Fis1, ROS, MDA, mitochondrial structural damage	A; AMPK/NRF2 blockade; Pre: Post-onset untested	Zhang et al[86]

Selected models, groups and findings are shown; ↑/↓ denote increases/decreases. Minimum effective dose/minimum effective concentration require dose-response support; otherwise not reported. Botanical names and medicinal parts follow authoritative databases and pharmacopoeial/European Medicines Agency monographs. Evidence strength summarizes model directness, mechanistic tests and timing relevance. Model categories: A: In vivo myocardial ischemia/reperfusion (I/R) in an intact animal; B: Isolated-heart I/R without intact systemic context; C: Cellular hypoxia/reoxygenation (R) or oxygen-glucose deprivation/R surrogates; D: Permanent myocardial ischemia/myocardial infarction (MI) without reperfusion; E: Other injury models. A/B directly model cardiac I/R; C models cellular components; D/E provide indirect evidence. Categories are not overall evidence-quality ranks. Assoc: No targeted causal test; named perturbations support the tested pathway, not necessarily fission mediation; comparators do not establish necessity. Pre: Before injury (post-onset initiation untested); Isch: During ischemia; Rep: At/after reperfusion onset. Acute-treatment relevance depends on start time, model, route and exposure; Pre + Rep does not establish Rep-only efficacy. Post-MI: After non-reperfused MI (post-infarction recovery). Cellular timing does not establish clinical windows; timing relevance does not establish clinical efficacy. Sample sizes are endpoint-specific and per group unless independent experiments are specified; cellular n retains the reported replication unit. Only explicitly reported outcome values are shown; graph estimates are excluded. ≈ denotes an approximation stated by the source authors; dispersion is retained as reported. Tongmai Yangxin Pill timing is inconsistently reported in Yu et al[84], precluding a firm timing-relevance assessment. R: Randomization; B: Reported blinding stage; yes: Reported; NR: Not reported; GRS: Ginsenoside Rb1-ruscogenin-schisandrin combination; LAD: Left anterior descending coronary artery; I/R: Ischemia/reperfusion; H/R: Hypoxia/reoxygenation; Drp1: Dynamin-related protein 1; AMPK: AMP-activated protein kinase; NRVMs: Neonatal rat ventricular myocytes; MED: Minimum effective dose; MEC: Minimum effective concentration; EF: Ejection fraction; Bcl-2/Bax: B-cell lymphoma 2 to Bcl-2-associated X protein; LDH: Lactate dehydrogenase; AMPK: AMP-activated protein kinase; PBM: Proprietary botanical medicine; MI: Myocardial infarction; LAD: Left anterior descending coronary artery; SMI: Shenmai Injection; FS: Fractional shortening; LC3-II: Light chain 3-II; ΔΨm: Mitochondrial membrane potential; Fis1: Mitochondrial fission 1 protein; mPTP: Mitochondrial permeability transition pore; CRF: Chronic renal failure; GXN: Guanxinning Injection; MFN1: Mitofusin 1; OPA1: Optic atrophy 1; cTnT: Cardiac troponin T; CK-MB: Creatine kinase-MB; CMECs: Cardiac microvascular endothelial cells; OGD/R: Oxygen-glucose deprivation/reoxygenation; SSNX: Shuangshen Ningxin Formula; NR4A1: Nuclear receptor subfamily 4 group A member 1; Mff: Mitochondrial fission factor; HK II: Hexokinase II; Cyt c: Cytochrome c; THSWD: Taohong Siwu Decoction; LVESV: Left ventricular end-systolic volume; ROS: Reactive oxygen species; Akt: Protein kinase B; QSYQ: Qishen Yiqi Dripping Pill; LVEF: Left ventricular ejection fraction; PGC-1α: Peroxisome proliferator-activated receptor-γ coactivator 1α; TFAM: Mitochondrial transcription factor A; DQ: Danqi Soft Capsule; TMYX: Tongmai Yangxin Pill; cTnI: Cardiac troponin I; ERα: Estrogen receptor α; PC6: Pericardium 6, Neiguan; ST36: Stomach 36, Zusanli; CV4: Conception vessel 4, Guanyuan; EA: Electroacupuncture; SOD: Superoxide dismutase; MDA: Malondialdehyde; NRF2: Nuclear factor erythroid 2-related factor 2.