Systematic Review of Preclinical Evidence on Parenteral Administration of Schizonepeta Tenuifolia Sole Extract

Article information

Perspect Integr Med. 2025;4(2):76-86

Publication date (electronic) : 2025 June 30

doi : https://doi.org/10.56986/pim.2025.06.002

Chaeheon Lee ¹^,^†

, Jihwan Choi ¹^,^†

, Ji Eun Choo ²

, Ga Ram Yang ²^,³^,⁴

, Hyung-Cheal Mun ⁵

, Won Gun An ³

, Cheol-Hyun Kim ⁶

, Jaehyo Kim ⁷

, Sangkwan Lee ⁶

, Hongmin Chu ⁶^,⁸^,

¹Department of Korean Medicine, Wonkwang University, Iksan, Republic of Korea

²Usher Bio Co., Ltd., Incheon, Republic of Korea

³Department of Pharmacology, College of Korean Medicine, Pusan National University, Busan, Republic of Korea

⁴Kangchoo Korean Medicine Clinic, Incheon, Republic of Korea

⁵Mokpo Beautiful Care Hospital, Mokpo, Republic of Korea

⁶Department of Internal Medicine and Neuroscience, College of Korean Medicine, Wonkwang University, Iksan, Republic of Korea

⁷Department of Meridian and Acupoint, College of Korean Medicine, Wonkwang University, Iksan, Republic of Korea

⁸MapoHongik Korean Medicine Clinic, Seoul, Republic of Korea

^*Corresponding author: Hongmin Chu, MapoHongik Korean Medicine Clinic, 16 Samgae-ro, Mapo-gu, Seoul 04173, Republic of Korea, Email: hongminchu2@gmail.com

†Two authors were contributed equally.

Received 2025 February 8; Revised 2025 May 15; Accepted 2025 May 28.

Abstract

While steroids effectively control inflammation, their long-term use causes severe side effects, necessitating safer alternatives. Schizonepeta tenuifolia (S. tenuifolia) from the Lamiaceae family demonstrates potential as a natural therapeutic option through its anti-inflammatory and antiviral properties. This systematic review analyzed in vivo and in vitro studies of S. tenuifolia extract, focusing on non-oral administration routes to evaluate its therapeutic potential (n = 13). The extract effectively inhibited nuclear factor kappa-light-chain-enhancer of activated B cells and mitogen-activated protein kinase pathways, reduced inflammatory cytokine production, and showed antiviral effects. Additional benefits include wound healing and antiplatelet activity which enables targeted inflammation control without systemic immune suppression. The extract showed promise in conditions requiring targeted pathway modulation, such as inflammatory conditions needing selective cytokine inhibition, viral infections where interferon modulation is beneficial, and disorders with dysregulated mitogen-activated protein kinase signaling. These effects were achieved without the systemic immunosuppression typical of steroid treatments. While S. tenuifolia extract could serve as a safer alternative through non-oral administration routes, further research is needed to optimize extraction methods, identify key marker compounds, and determine optimal administration routes including topical administration and injectable formulations.

Keywords: anti-inflammatory agents; antiviral agents; herbal medicine; mitogen-activated protein kinase; steroid

Visual abstract

Introduction

Glucocorticoid steroids are widely used in the medical field for their strong anti-inflammatory and immunosuppressive effects, which can quickly alleviate inflammation [1]. However, the long-term use of steroids can lead to side effects such as muscle atrophy, osteonecrosis, osteoporosis, hypertension, hormonal changes, and metabolic changes [2]. Degenerative changes due to muscle atrophy can result in the infiltration of adipose tissue into the muscles [3]. Glucocorticoid-induced skeletal muscle atrophy has been shown to occur by enhancing protein degradation pathways and inhibiting protein synthesis, particularly affecting fast-twitch muscle fibers [4]. In addition, extended steroid use can induce adrenal suppression, and when steroid dosage is reduced in such cases it may cause symptoms like fatigue, joint pain, muscle pain, and hypotension [2]. In contrast, natural substance extracts have garnered attention for their relatively safe profile and minimal side effects compared with steroid medications. Recent studies have demonstrated that certain natural products and their components can effectively mitigate inflammation [5,6]. Consequently, researchers are actively exploring the application of injectable agents containing natural extracts for treating localized inflammation. Natural products comprise numerous compounds with anti-inflammatory properties, with plants from the Labiatae (Lamiaceae) family being particularly noteworthy.

Their anti-inflammatory effects are mediated through various mechanisms, including nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway inhibition, proinflammatory cytokine production and cyclooxygenase-2, and inducible nitric oxide synthase expression suppression [7]. Among the Lamiaceae family, Schizonepeta tenuifolia (S. tenuifolia) is distinguished for its notable anti-inflammatory, antiallergic, and antiviral properties [8]. This annual herbaceous plant, belonging to the Schizonepeta genus, has been traditionally used to treat colds, fever, and headaches [9]. Most S. tenuifolia-based herbal medicines are primarily administered orally, and clinical studies have predominantly focused on decoction-based complex formulations [10]. Some studies have investigated components of S. tenuifolia, but a systematic review of preclinical studies has not been reported [8].

To this end, a systematic review of in vivo and in vitro studies on S. tenuifolia was conducted to explore its potential for developing non-oral administration routes, including injectable and topical applications. While injections were the focus, studies using topical agents were included to broaden the understanding of local anti-inflammatory and antiviral effects relevant to non-oral therapy development. Studies using S. tenuifolia (Jingjie) alone, which were relevant in vivo and in vitro research, with a focus on developing formulations that can be administered through non-oral methods were systematically reviewed. The goal of this study was to review preclinical research on S. tenuifolia and to highlight its potentially active components and key biological pathways.

Materials and Methods

1. Study registration

The protocol for this systematic review was registered with the Open Science Framework [11], and the study followed the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines [12].

2. Database and search strategy

In this study, the search strategy included English, Chinese, and Korean databases [MEDLINE (https://pubmed.ncbi.nlm.nih.gov/), EMBASE (https://www.embase.com), Cochrane (https://www.cochranelibrary.com/), CNKI (https://www.cnki.net/index/), AMED (https://web-p-ebscohost-com), Riss (https://www.riss.kr/index), and OASIS (https://oasis.kiom.re.kr/index)]. The main keywords included “Schizonepeta tenuifolia (S. tenuifolia),” “extract, injection, administration,” and “animal, human clinical trial” (Supplementary Material 1).

3. Study selection process

This study mainly focused on mechanisms of action and the effects of S. tenuifolia extract when administered through non-oral routes. Therefore, animal experiments involving oral administration of S. tenuifolia extract and studies on combined extracts of S. tenuifolia were excluded from this review. However, we included studies of combined extracts administered through non-oral route, if they provided separate data for S. tenuifolia extract alone. In addition, studies utilizing S. tenuifolia extract alone as a topical agent were also included to comprehensively assess the local effects and therapeutic potential of S. tenuifolia extract. Although the absorption and pharmacokinetic properties differ between topical and injectable routes, topical studies offer relevant insights into local anti-inflammatory activity and tissue interaction, which are valuable for the early-phase exploration of non-oral formulations.

4. Data extraction process

After importing all studies into a reference management tool (Zotero, developed by the Roy Rosenzweig, maintained by Corporation for Digital Scholarship, Version 7), duplicates were removed as the first step of the process. Then, using Microsoft Excel, the extracted papers were listed, and 2 authors (HC, JEC) reviewed the titles and abstracts to determine eligibility during the initial search conducted from August 2 to August 17, 2024. The studies initially selected were further reviewed in full text by 2 other authors (CL, JC) from August 24 to September 3, 2024. In cases where there was a disagreement between the 2 researchers and they could not reach a consensus, a 3^rd party (GRY), who was experienced in chemical life engineering and Korean medicine, reviewed the full text to make the final decision. There were no restrictions on language, publication date, or publication status.

Results

1. Search results

The primary search identified a total of 239 studies (Figure 1). Clinical trials, RCTs, reviews, letters, and other studies unrelated to the preclinical research on S. tenuifolia were excluded by reviewing the titles and abstracts. In addition, botanical papers on the growth and origin of S. tenuifolia were also excluded. From this initial extraction, 35 papers were selected. Upon full-text review, duplicate studies and oral administration studies were removed, resulting in a final selection of 13 studies for review [13–25].

Figure 1

Flowchart of the systematic review.

The review began with 239 studies identified via database searches. Exclusion criteria were clinical trials, RCTs, reviews, letters, unrelated papers, and botanical studies on S. tenuifolia growth/origin. This narrowed the results to 35 articles. Full-text review excluded duplicates and studies on oral administration, yielding a final 13 papers that met all criteria. The flowchart visually details the screening process, showing the number of studies included or excluded at each step, from initial identification to final selection for analysis.

2. Categorized method and study characteristics

The studies were categorized into cell-based experiments (in vitro) and animal experiments (in vivo) and then organized into 4 sections: (1) Data from in vivo studies; (2) In vivo study outcome data; (3) General characteristics of in vivo studies; and (4) In vitro study outcome data. For studies that conducted both in vivo and in vitro research, information was recorded in duplicate. The characteristics and results of the included in vivo studies are presented in Table 1, and in vitro studies in Table 2. Additional characteristics of the in vivo studies are provided in the Supplementary files.

Table 1

In Vivo Studies of Non-Oral Schizonepeta Tenuifolia Extract

Table 2

In Vitro Studies of Non-Oral Schizonepeta Tenuifolia Extract

2.1. In vivo study

The animal models utilized in these studies exclusively employed mice and rats. The diverse range of models included wound healing, EV71-induced infection, lipopolysaccharide (LPS)-induced bone erosion, tail bleeding, 2,4-dinitrochlorobenzene -induced atopic dermatitis, compound 48/80-induced systemic anaphylaxis, and immunoglobulin E-mediated passive cutaneous anaphylaxis models. This variety underscores the multifaceted potential therapeutic applications of S. tenuifolia extract. Additional characteristics of in vivo studies are summarized in Supplementary Material 2.

2.2. In vitro study

In vitro studies selected for this review were conducted in various East Asian countries: 8 from Korea, 1 from Japan, 1 from China, and 1 from Taiwan. Extraction methods for S. tenuifolia varied across the studies, with 4 using distilled water, 3 using ethanol, and 3 using methanol, highlighting the importance of extraction technique in phytochemical research.

In vitro studies, evaluated numerous parameters including cell viability, inflammatory mediator production, messenger ribonucleic acid (mRNA) expression, gene expression, protein phosphorylation, and various functional assays (Table 2 and Supplementary Material 3). Results consistently demonstrated anti-inflammatory, antiviral, and anti-allergic effects of S. tenuifolia extract across multiple cell types. The underlying mechanisms primarily involved inhibition of NF-κB and mitogen-activated protein kinase (MAPK) signaling pathways, and modulation of cytokine production. Additional observed effects included promotion of wound healing, inhibition of platelet aggregation, and suppression of osteoclast differentiation and bone resorption.

Discussion

A systematic review of preclinical studies of S. tenuifolia, when administered through non-oral routes, was conducted to explore the mechanisms of action. The main pathways identified through this review were as follows: inhibition of the NF-κB and MAPK pathways leading to a decrease in inflammatory cytokines, and the antiviral effect through the suppression of the ROS-p38 MAPK-hnRNP A1 axis. This review aimed to narrow down the key target pathways for future drug development based on S. tenuifolia extract.

This study serves as foundational research exploring S. tenuifolia extract as a potential candidate for the development of natural injectable therapeutics and investigates its mechanisms of action. While steroids have strong anti-inflammatory effects, they also exert a broad impact on the immune system. Despite their potent anti-inflammatory properties, steroids are unsuitable for long-term use due to their extensive immunosuppressive effects, and influence on multiple bodily systems [26]. In addition, steroids activate the ubiquitin proteasome system which is associated with protein degradation, promoting muscle breakdown and muscle atrophy [2]. There are case reports about acute muscle atrophy induced by steroid use [27,28]. In a study by Boonen et al [27], it was reported that Cushing’s syndrome and pelvic girdle muscle weakness caused by a single epidural triamcinolone injection. In a case reported by Jordan et al [28], a shoulder joint injection with triamcinolone led to weakness in both lower limbs after 18 hours. Furthermore, long-term use of steroids can induce tolerance, resulting in the need for higher doses [2]. This may increase the possibility of more severe side effects. Moreover, discontinuing steroid use may cause rebound inflammation in which the inflammation returns or worsens. Thus, many patients find it difficult to stop taking steroids, and this leads to chronic usage [29].

Natural substances, like S. tenuifolia extract, selectively inhibit the NF-κB and MAPK pathways, and these pathways have a more targeted effect on inflammation control rather than causing systemic immune suppression [30]. Therefore, it is considered that these natural substances can regulate inflammation without affecting the entire immune system, targeting only specific inflammation-related pathways, which results in relatively fewer side effects than glucocorticoids [31].

S. tenuifolia exhibits anti-inflammatory properties through several mechanisms: it inhibits MAPK phosphorylation, NF-κB activation, inflammatory cytokine production, and the expression of cyclooxygenase-2 and inducible nitric oxide synthase [9,32]. The primary chemical component of S. tenuifolia is its volatile oil which contains a variety of compounds, including flavonoids, and glycosides. In addition, the volatile oil is composed mainly of terpenoids, aldehydes, ketones, quinones, alcohols, phenols, esters, carboxylic acids, and alkenes [8,33–35]. Research by Choi et al [18] showed its effectiveness against atopic dermatitis in mice by suppressing mast cell degranulation, and modulating proinflammatory cytokine production and inflammatory pathways such as NF-κB and MAPK signaling pathways. The antiallergic effects of S. tenuifolia are also noteworthy [19,36]. In a study by Lin et al [36], it was reported that Schizonepeta tenuifolia exerted antiallergic effects by suppressing degranulation and regulating cytokine production in IgE-stimulated RBL-2H3 mast cells, notably decreasing pro-inflammatory cytokines such as IL-4, IL-6, IL-13, TNF-α, and IFN-γ while enhancing the anti-inflammatory cytokine IL-10, through modulation of immune responses rather than cytotoxicity. Another study [19] revealed the capacity of the extract to prevent compound 48/80-induced systemic allergic reactions and block histamine release from mast cells. In a study by Qin et al [37], it was reported that essential oil from S. tenuifolia exerted antidepressant effects in an LPS-induced mouse model by reducing IL-1β–mediated neuroinflammation through multi-target and multi-pathway mechanisms. The neuroprotective effects of S. tenuifolia are also notable. In a study by Yoon et al [38], its methanolic extract was reported to protect neuronal cells from hydrogen peroxide-induced cytotoxicity by mitigating oxidative DNA damage.

Research on the antiaging effects of S. tenuifolia is ongoing. In a study by Gu et al [39], it was reported that S. tenuifolia ethanolic extract alleviated UVB-induced photoaging in hairless mice by reducing wrinkle formation and skin dehydration through inhibition of MAPK and RAGE signaling pathways. These wide-ranging therapeutic effects stem from key components of S. tenuifolia, with its volatile oil being predominantly composed of (+)-pulegone and (−)-menthone, which have been strongly correlated with its antioxidant activity [40].

The anti-inflammatory properties and inhibitory mechanisms of S. tenuifolia extracts have been extensively documented through various in vivo and in vitro experimental models. Components underlying S. tenuifolia’s therapeutic effects include immunoregulation of MAPK and NF-κB pathways, and expression of Th1- and Th2-related inflammatory cytokines. A study by Byun [23], it was demonstrated that ethanol extraction of S. tenuifolia suppressed the production of nitric oxide, prostaglandin E2, interleukin (IL)-1β, and rosis factor alpha (TNF-α) in LPS-stimulated bone marrow-derived macrophages by inhibiting NF-κB and MAPK pathways. This anti-inflammatory activity involved the inhibition of multiple components: inhibitor of NF-κB alpha (IκBα), MAPK phosphorylation (p38, ERK1/2, JNK), and p65 nuclear translocation. In a study by Kang et al [20], it was reported that water extraction of S. tenuifolia reduced interferon (IFN)-γ and IL-4 levels while increasing IL-2 levels in peripheral blood mononuclear cells and splenocytes. This may be linked to enhanced nuclear translocation of nuclear factor of activated T cells cytoplasmic 2, and reduced nuclear translocation of p65, as demonstrated in their research on splenocytes [20]. In a study by Kang et al [21], it was reported that in LPS-treated peritoneal macrophages, water extraction of S. tenuifolia inhibited both IκBα degradation, and JNK/stress-activated protein kinase activation, consequently reducing the production of TNF-α and IL-6. In a study by Choi et al [18], it was reported that water extraction of S. tenuifolia was effective in reducing inflammatory markers in atopic dermatitis, demonstrated by decreased immunoglobulin E, TNF-α, and IL-6 levels in a 2,4-dinitrochlorobenzene-induced mouse model. In a study by Sohn et al [22], both S. tenuifolia extract and its purified chemicals (rosmarinic acid, pulegone, and 2α,3α,24-trih ydroxy olean-12 en-28 oic acid) were shown to inhibit gene expression in human mast cell-1 cell line stimulated with phorbol 12-myristate 13-acetate plus the antibiotic ionophore A23187. The affected genes were those involved in toll-like receptor and MAPK pathways, apoptosis processes, cytokine-cytokine receptor interactions, and p53 signaling pathways. S. tenuifolia appears to reduce inflammation by regulating multiple signaling pathways, especially NF-κB and MAPK, which results in decreased proinflammatory mediator production.

Several studies [14,16,24] have also investigated the anti-viral mechanisms of S. tenuifolia. In a study by Chen et al [14], it was reported that S. tenuifolia inhibits enterovirus 71 replication by suppressing the reactive oxygen species (ROS)–p38 MAPK–heterogeneous nuclear ribonucleoproteins (hnRNP) A1 axis. Their study demonstrated that S. tenuifolia maintained cap-dependent translation by preserving eukaryotic initiation factor 4G activity and inhibits IRES-dependent translation by preventing hnRNP A1 translocation to the cytoplasm. In addition, water extraction of S. tenuifolia exhibited antiviral activity against enterovirus 71 involved multiple mechanisms: reducing viral attachment and entry, preventing eukaryotic initiation factor 4G cleavage by EV71 protease 2Apro, suppressing virus-induced reactive oxygen species (ROS) formation, and hindering the relocation of hnRNP A1 from the nucleus to the cytoplasm. This process was accompanied by a decline in EV71-associated hyperphosphorylation of both p38 kinase and epidermal growth factor receptor pathway substrate 15 [14]. In a study by Ng et al [24], S. tenuifolia’s antiviral properties against noroviruses were further explored. In RAW 264.7 cells infected with murine norovirus 1, the methanol extraction of S. tenuifolia increased IRF3 phosphorylation, leading to elevated murine IFN-β and reduced viral RNA levels [24]. Similar effects were observed in the experiment using human norovirus Norwalk virus replicon-harboring cell (HG23 cells), where the extract increased both IFN-β and IFN-γ mRNA levels, resulting in decreased viral RNA [24]. Interestingly, in a study by Liu et al [16], it was reported that charred S. tenuifolia (SSC) exhibited more potent antiviral activities compared to raw S. tenuifolia (raw SS). In their cell-based study, SSC demonstrated not only hemostatic effects but also stronger antiviral effects against RSV compared to raw SS [16]. This suggests potential modifications in the chemical structure during the stir-frying process that could enhance its antiviral properties. Additionally, when treated with LPS, carbonized S. tenuifolia resulted in a dose-dependent decrease in NO, IL-6, TNF-α, and IL-1beta. In contrast, ethanol extraction from raw S. tenuifolia led to an increase in IL-6 and TNF-α, highlighting a unique difference in the bioactivity of the carbonized versus raw forms.

In a different therapeutic context, in a study by Kim et al [15], S. tenuifolia’s effects on osteoclasts was investigated. They observed that S. tenuifolia prevented bone loss by suppressing osteoclast development and function by inhibiting Akt and IκB phosphorylation and downregulating nuclear factor of activated T cells, cytoplasmic 1 and c-fos expression, ultimately leading to enhanced bone mineral density [15]. This finding indicates potential therapeutic benefits of S. tenuifolia for osteoporosis treatment.

In an in vitro and in vivo study by Isohama et al [13], it was reported that S. tenuifolia had a role in wound healing. Their research with DJM-1 cells showed that the methanol extraction of S. tenuifolia increased both aquaporin 3 mRNA expression and protein levels, even when TNF-α was present. The increased aquaporin-3 expression led to enhanced migration activity of DJM-1 cells following treatment with methanol extraction of S. tenuifolia. The in vivo experiment on hairless mice supports these in vitro findings, confirming the extract’s capacity to accelerate wound healing [13].

Regarding platelet function, in a study by Jeon et al [25], it was reported that antiplatelet effects of methanol extraction of S. tenuifolia was exerted by reducing phosphorylation of ERK, JNK, MEK, and Akt, therefore inhibiting MAPK and PI3K/Akt pathways. Collagen-induced platelet aggregation, intercellular Ca²⁺ level, adenosine triphosphate secretion, and fibrinogen-binding to integrin αIIbβ3 all decreased when platelets were treated with methanol extraction of S. tenuifolia [25]. In a study by Shin et al [19], it was reported that water extraction of S. tenuifolia could inhibit histamine release in rat peritoneal mast cells, likely through membrane stabilization or interference with signaling pathways.

In conclusion, S. tenuifolia exerts its therapeutic potential through multi-faceted mechanisms. Its primary effects include inhibition of inflammatory pathways (NF-κB and MAPK), immune response modulation, and antiviral activities. Additional effects of S. tenuifolia extract include antiplatelet effect, aquaporin-3 upregulation for wound healing, and histamine release inhibition. These findings suggest that S. tenuifolia, when administered through various routes, and extracted in various ways, can act through diverse mechanisms targeting inflammation, viral infections, and bone disorders. S. tenuifolia holds significant potential for treating various conditions. The summary of the therapeutic effects is presented in (Figure 2).

Figure 2

Summary of therapeutic effects of S. tenuifolia.

S. tenuifolia extract exerts therapeutic effects via several pathways: (1) Anti-inflammatory and immunomodulatory: inhibits NF-κB and MAPK, downregulates mediators (NO, iNOS, COX-2, PGE2), cytokines (TNF-α, IL-1β, IL-6, IL-8), and modulates Th1/Th2 balance, related factors, and production; (2) Antiviral: suppresses replication via ROS-p38 MAPK-hnRNPA1, prevents eIF4G cleavage, enhances interferons, and resistance to norovirus/respiratory viruses; (3) Other: promotes wound healing, inhibits osteoclastogenesis, suppresses histamine, and has anti-platelet effects via MAPK/Akt.

COX-2 = cyclooxygenase-2; eIF4G = eukaryotic initiation factor 4G; hnRNP A1 = heterogeneous nuclear ribonucleoprotein A1; MAPK = mitogen-activated protein kinases; NO = nitric oxide; iNOS = inducible nitric oxide synthase; PGE2 = prostaglandin E2; ROS = reactive oxygen species.

It is important to determine whether the injectable formulation of S. tenuifolia should be based on a water, methanol or ethanol extraction, and whether the extract should be raw or carbonized. Selection of a suitable main marker compound whether a single or multiple markers is also required. In addition, the administration route needs to be carefully planned, deciding whether to apply the extract as a localized topical treatment or via intramuscular injection, or to develop it for intravenous injection to act systemically. For ease of storage, we propose initially developing a powder formulation that can be reconstituted in saline or dextrose water for topical application or intramuscular injection. After confirming safety, intravenous formulations may be explored. This review aimed to identify key marker compounds and mechanisms of action for injectable formulations of S. tenuifolia. MAPK signaling and antiviral activity emerged as major pathways. In addition, the purpose of this review to provide a comprehensive synthesis of the existing literature rather than to perform quantitative analyses. A meta-analysis was not feasible due to the limited number of comparable studies but this may be possible in the future as more data become available.

This study has the strength of systematically reviewing a wide range of in vivo and in vitro studies to explore the potential use of S. tenuifolia as a non-oral route medication. In addition, the inclusion of English, Chinese, and Korean databases in the search to retrieve studies is another advantage, and enhances the review’s scope. Since many studies included both in vivo and in vitro experiments, this research provided a comprehensive overview of preclinical studies. The key mechanisms of S. tenuifolia were identified as the inhibition of MAPK, NF-κB pathways, and inflammatory cytokines related to Th1- and Th2 responses. Among the included studies, Zhang et al [17] reported that the combination of Yizhiren and Schizonepeta outperformed the standard drug, dexamethasone, as an external treatment for atopic dermatitis, showing no hepatotoxicity in atopic-induced rats. In addition, in a study by Liu et al [16], it was demonstrated that carbonized Schizonepeta had greater hemostatic and antiviral effects compared with its raw form, suggesting the potential use of carbonized Schizonepeta for future drug development.

However, this study has several limitations. While the systematic review was conducted with the aim of identifying an injectable drug, no studies were found that involved intravenous injection of S. tenuifolia extract; the majority of studies focused on local injection or topical application. Therefore, further research is needed if Schizonepeta extract is to be used via intravenous administration. In addition, there were no studies that isolated specific components, through hot water or methanol extraction, of S. tenuifolia, and conducted animal experiments to develop Schizonepeta into a natural pharmaceutical. Schizonepeta is known to contain components like limonene and thymol. While this review identified mechanisms related to anti-inflammatory effects, immune modulation, and antiviral activity, further research is necessary to explore its antihistamine effects. In addition, to assess the methodological quality of the included studies, the Systematic Review Centre for Laboratory animal Experimentation (SYRCLE)’s Risk of Bias tool was applied for in vivo studies, and Modified SYRCLE’s Risk of Bias tool was applied for in vitro studies. The evaluation indicated that reporting standards varied among the studies, with several domains assessed as “unclear” due to insufficient methodological detail. While this reflects common challenges in synthesizing preclinical evidence, careful interpretation of the findings is warranted (the summarized results are provided in Figures 3 and 4. Full evaluation criteria of the Modified SYRCLE’s risk of bias tool are available in Supplementary Material 4). Finally, despite an extensive systematic literature review being conducted, some relevant studies may have been overlooked due to database limitations, posing another limitation of this research.

Figure 3

SYRCLE’s Risk of Bias of in vivo studies.

Figure 4

Modified SYRCLE’s Risk of Bias of in vitro studies.

Conclusion

This systematic review examined preclinical investigations, encompassing both in vivo and in vitro studies, to assess S. tenuifolia’s therapeutic potential via non-oral administration routes. The findings demonstrated significant anti-inflammatory, antiviral, and immunomodulatory properties, with molecular mechanisms primarily involving the suppression of MAPK and NF-κB pathways, alongside the modulation of Th1- and Th2-mediated inflammatory cytokine production. The superior safety profile of Schizonepeta, compared to standard steroid treatments, and the enhanced antiviral effects of carbonized forms, indicated its potential for future drug development. However, the review revealed several limitations, including the lack of studies on intravenous administration routes, and the absence of research on component isolation of Schizonepeta for targeted drug development. In addition, further studies are needed to explore its antihistamine properties and to develop injectable formulations. Despite these limitations, this comprehensive review serves as a foundational step for future research, underscoring the promising therapeutic applications of S. tenuifolia as a safer alternative to steroid-based anti-inflammatory treatments.

Supplementary Materials

Supplementary materials are available at doi: https://doi.org/10.56986/pim.2025.06.002

pim-2025-06-002-Supplementary-1.docx

pim-2025-06-002-Supplementary-2.docx

pim-2025-06-002-Supplementary-3.docx

pim-2025-06-002-Supplementary-4.docx

Notes

Author Contributions

Conceptualization: HC, JEC, CL, and JC. Methodology: HC and JEC. Software: HC and JEC. Formal investigation: HC. Resources: HC. Data Analysis: HC and GRY. Writing of the Original Draft: HC, CL, and JC. Review and Editing: CL and JC.

Conflicts of Interest

The authors have no conflicts of interest to declare.

Author Use of AI Tools Statement

The authors used ChatGPT (OpenAI) solely for grammar and language improvement during the revision of this manuscript. The AI tool did not contribute to the scientific content, data analysis, or interpretation. The authors take full responsibility for all content.

Funding

This research received support from a grant provided by the Korea Health Technology R&D Project through the Korea Health Industry Development Institute, funded by the Ministry of Health and Welfare, Republic of Korea (grant no.: HF20C0113).

Ethical Statement

Not applicable.

Data Availability

All relevant data are included in this manuscript.

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Article information Continued

This is an open access article under the CC BY-NC license (http://creativecommons.org/licenses/by-nc/4.0/).

Authors, year, country, [reference]	Therapeutic effect	Group number (N, group)	Observation period	Route	Dose	Measured parameter and results (vs. control group)
Isohama et al, 2014, Japan [13]	Wound healing	2, 8	7 d observation & treatment	Topical	Ointment containing 5 μg/site of STE. Treated once daily	↓: Initial wound area
Chen et al, 2017, Taiwan [14]	Antiviral (EV71)	2, 15–16	14 d treatment, 17 d observation	Intraperitoneal	250 mg/kg	↑: Survival rate (87.5%) ↓: Severity & progression of neurologic symptoms
Kim et al, 2016, Korea [15]	Anti-inflammatory/bone Health	3, 5	8 d treatment & observation	LPS: Intraperitoneal	LPS (5 mg/kg)	↑: BV/TV (15%), Tb.N (20%) ↓: Tb.Sp (15%), TRAP-positive osteoclasts
Zhang et al, 2020, Korea [17]	Atopic dermatitis	6, 8	5 wks	Topical application on dorsal skin	200 μL of 30% STB, 30% AOM, or mix, in BG; applied twice daily	↓: Skin thickness, Mast cell, Serum IgE, cytokines
Choi et al, 2012, Korea [18]	Atopic dermatitis	3, 5	17 d	Intraperitoneal	250 mg/kg	↓: Epidermal thickness (38.15%) IgE serum level (46.25%), TNF-α (41.97%), IL-6 (70.42%) Protein expression of NF-κB and MAPKs (JNK, ERK, p38)
Shin et al, 1999, Korea [19]	Anti-allergic/anaphylaxis	Multiple, at least 3	1 h for systemic anaphylaxis	Intraperitoneal	STAE 0.005 to 1 g/kg; compound 48/80	↓: Mortality rate, plasma histamine

Table 2

In Vitro Studies of Non-Oral Schizonepeta Tenuifolia Extract

Authors, year, country, [reference]	Therapeutic effect/key Finding	Extraction method/solvent	Assay type/treatment conditions	Key results (quantitative/qualitative)
Kang et al, 2010, Korea [20]	Immunomodulatory/anti-inflammatory	Distilled water	Splenocytes /PBMCs + anti-CD3± STE/DEX; Splenocytes + anti-CD3/IL-12 +STE/CsA /DE	No cytotoxicity (up to 100 μg/mL) ↑: Mitogenic activity (Splenocytes) (1.5–2.5 fold), IL-2 transcription, IFN-γ transcription (8 h, 24 h), IL-2 (Splenocytes /PBMCs), NFATc2 nuclear translocation, STAT4 phosphorylation, STAT6 phosphorylation ↓: IFN-γ transcription (at 48 h), IL-4 transcription, IFN-γ & IL-4 (splenocytes /PBMCs), p65 nuclear translocation
Kang et al, 2010, Korea [21]	Anti-inflammatory	Distilled water	Macrophages + LPS ± STE (50–200 μg/mL) / DE	No cytotoxicity (up to 400 μg/mL) ↓: TNF-α (6 h, 24 h), IL-6 (6 h), TNF-α mRNA (4 h), IkBα degradation (15 min), p65 nuclear translocation, JNK/SAPK phosphorylation, c-Jun phosphorylation, ATF-2 phosphorylation
Sohn et al, 2012, Korea [22]	Anti-inflammatory/gene regulation	Unidentified	HMC-1 + PMA/A23187 ± ST/chemicals	No cytotoxic effect (STE only) ↑: Viability (HMC-1 stimulated) (specific chemicals) ↓: TNF-α, IL-6, IL-8 (specific chemicals), specific gene expression (8 genes)
Byun et al, 2014, Korea [23]	Anti-inflammatory	70% ethanol	BMDMs + LPS ± STE (25–100 μg/mL	Cell viability: maintained (up to 100μg/mL, 24h) ↓: NO, iNOS, COX-2, PGE2, IL-1β, IL-6, CD80, CD86, TNF-α, TLR4 protein, p38 phosphorylation, ERK1/2 phosphorylation, IkB-α phosphorylation, p65 nuclear translocation
Isohama et al, 2014, Japan [13]	Skin barrier function/wound healing	Methanol	DJM-1 ± TNF-α +STE/DEX; Scratched DJM-1 + STE ± AQP3 siRNA	↓: AQP3 mRNA & protein (TNF-α treated), water permeability ↑: AQP3 mRNA (STE only) (up to 2.5-fold), AQP3 protein (STE only), AQP3 mRNA (TNF-α + STE), gap closure/migration (effect abolished by AQP3 siRNA)
Ng et al, 2018, Korea [24]	Antiviral (norovirus)	80% methanol	HG23 (replicon) ± STE/RBV /LGEO/chemicals; RAW + MNV-1 ± STE/RB	No cytotoxicity (HG23 cells) ↓: Viral RNA (HG23), MNV-1 plaque formation, MNV-1 RNA levels ↑: IFN-β mRNA (HG23), IFN-γ mRNA (HG23), mIFN-β mRNA (RAW 264.7), IRF3 phosphorylation (RAW 264.7)
Jeon et al, 2019, Korea [25]	Anti-platelet aggregation	Methanol	Washed platelets + collagen/ADP/thrombin ± STE (vehicle control)	↓: Collagen-induced platelet aggregation, [Ca²⁺]i, ATP secretion, fibrinogen binding to integrin αIIbβ3, Phosphorylation (ERK, JNK, MEK, Akt) No effect: ADP-induced aggregation
Chen et al, 2017, Taiwan [14]	Antiviral (EV71)	Distilled water	RD/Vero cells + EV71 ± STE; + p38 MAPK Inhibitor (SB202190); +H₂O₂; Transfection	↓: EV71 plaque formation, RNA replication, Protein synthesis, Viral particle production, IRES-dependent translation, eIF4G cleavage, hnRNP A1 cytoplasmic translocation, p38 MAPK activation, EPS15 phosphorylation, EV71-induced ROS generation
Kim et al, 2016, Korea [15]	Anti-osteoclastogenesis/bone health	95% ethanol	BMMs + M-CSF + RANKL ± EEST; retroviral transfection	No cytotoxicity ↓: RANKL-induced osteoclast formation, Akt phosphorylation, IkB phosphorylation, c-Fos expression, NFATc1 expression, F-actin ring formation, bone resorption activity, osteoclast-specific gene expression (multiple genes)
Liu et al, 2021, China [16]	Anti-inflammatory/antiviral (RSV)	75% ethanol (v/v) for SS and SSC	RAW 264.7 + LPS ± SS/SSC; HEp-2 + RSV ± SS/SSC; chem. profile	↓: Anti-inflammatory effect (SSC vs SS), essential oil yield (SSC vs SS) (91%) ↑: Antiviral activity (SSC vs SS) Altered: chemical profile (charred vs raw)
Shin et al, 1999, Korea [19]	Anti-allergic/mast cell degranulation	Distilled water	Mast cells + Cpd 48/80 or anti-DNP IgE + DNP-HAS ± STA	↓: Histamine release ↑: TNF-α production

BMDMs = bone marrow-derived macrophages; bSS = β-Sitosterol; [Ca²⁺]i = intracellular calcium level; CO = caryophyllene oxide; COX-2 = cyclooxygenase-2; CPE = cytopathic effect; CsA = cyclosporine A; eIF4G = eukaryotic initiation factor 4G; EEST = ethanol extract of Schizonepeta tenuifolia; EPS15 = epidermal growth factor receptor pathway substrate 15; EV71 = Enterovirus 71; HG23 = human astrocytoma U-87 MG cells (infected with human enterovirus 71, strain HG23); hnRNP A1 = heterogeneous nuclear ribonucleoprotein A1; HP = hesperidin; IκB-α = inhibitor of kappa B alpha; IC50 = half-maximal inhibitory concentration; IFN-β = interferon-beta; IFN-γ = interferon-gamma; IL-1β = interleukin-1 beta; IL-2 = interleukin-2; IL-4 = interleukin-4; IL-8 = interleukin-8; iNOS = inducible nitric oxide synthase; IRES = internal ribosomal entry site; IRF3 = interferon regulatory factor 3; LEGO = Lonicera japonica-Erigeron annuus-Gleditsia sinensis extract; LG = luteolin-7-O-glucuroninide; MAPK = mitogen-activated protein kinases; MEK = mitogen-activated protein kinase kinase; mIFN-β = murine IFN-β; MNV-1 = murine norovirus 1; MTS assay = 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium assay; NFATc2 = nuclear factor of activated T-cells, cytoplasmic 2; NO = nitric oxide; PBMCs = peripheral blood mononuclear cells; PG = pulegone; PGE2 = prostaglandin E2; PI3K = phosphoinositide 3-kinase; pma = phorbol 12-myristate 13-acetate; (+)-M = (+)-Menthone; (−)-P = (S)-(−)-Pulegone; RA = rosmarinic acid; RANKL = receptor activator of nuclear factor kappa-B ligand; RBV = ribavirin; ROS = reactive oxygen species; RSV = respiratory syncytial virus; RT-qPCR = reverse transcription quantitative polymerase chain reaction; SI = selectivity index; SS = Schizonepeta tenuifolia raw; SSC = Schizonepeta tenuifolia charred; STAT4 = signal transducer and activator of transcription 4; STAT6 = signal transducer and activator of transcription 6; TH = 2α,3α,24-trihydroxyolean-12en-28oic acid; TLR4 = toll-like receptor 4; UA = ursolic acid; VP1 = viral protein 1; XTT assay = 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide assay.