Investigation of Anti-Liver Cancer Activity of the Herbal Drug FDY003 Using Network Pharmacology.

Globally, liver cancer (LC) is the sixth-most frequently occurring and the second-most fatal malignancy, responsible for 0.83 million deaths annually. Although the application of herbal drugs in cancer therapies has increased, their anti-LC activity and relevant mechanisms have not been fully studied from a systems perspective. To address these issues, we conducted a system-perspective network pharmacological investigation into the activity and mechanisms underlying the action of the herbal drug. FDY003 reduced the viability of human LC treatment. FDY003 reduced the viability of human LC cells and elevated their chemosensitivity. There were a total of 16 potential bioactive chemical components in FDY003 and they had 91 corresponding targets responsible for the pathological processes in LC. These FDY003 targets were functionally involved in regulating the survival, proliferation, apoptosis, and cell cycle of LC cells. Additionally, we found that FDY003 may target key signaling cascades connected to diverse LC pathological mechanisms, namely, PI3K-Akt, focal adhesion, IL-17, FoxO, MAPK, and TNF pathways. Overall, this study contributed to integrative mechanistic insights into the anti-LC potential of FDY003.

Exploration of the System-Level Mechanisms of the Herbal Drug FDY003 for Pancreatic Cancer Treatment: A Network Pharmacological Investigation.

Pancreatic cancer (PC) is the most lethal cancer with the lowest survival rate globally. Although the prescription of herbal drugs against PC is gaining increasing attention, their polypharmacological therapeutic mechanisms are yet to be fully understood. Based on network pharmacology, we explored the anti-PC properties and system-level mechanisms of the herbal drug FDY003. FDY003 decreased the viability of human PC cells and strengthened their chemosensitivity. Network pharmacological analysis of FDY003 indicated the presence of 16 active phytochemical components and 123 PC-related pharmacological targets. Functional enrichment analysis revealed that the PC-related targets of FDY003 participate in the regulation of cell growth and proliferation, cell cycle process, cell survival, and cell death. In addition, FDY003 was shown to target diverse key pathways associated with PC pathophysiology, namely, the PIK3-Akt, MAPK, FoxO, focal adhesion, TNF, p53, HIF-1, and Ras pathways. Our network pharmacological findings advance the mechanistic understanding of the anti-PC properties of FDY003 from a system perspective.

A Network Pharmacology Study on the Molecular Mechanisms of FDY003 for Breast Cancer Treatment.

Herbal medicines have drawn considerable attention with regard to their potential applications in breast cancer (BC) treatment, a frequently diagnosed malignant disease, considering their anticancer efficacy with relatively less adverse effects. However, their mechanisms of systemic action have not been understood comprehensively. Based on network pharmacology approaches, we attempted to unveil the mechanisms of FDY003, an herbal drug comprised of Lonicera japonica Thunberg, Artemisia capillaris Thunberg, and Cordyceps militaris, against BC at a systemic level. We found that FDY003 exhibited pharmacological effects on human BC cells. Subsequently, detailed data regarding the biochemical components contained in FDY003 were obtained from comprehensive herbal medicine-related databases, including TCMSP and CancerHSP. By evaluating their pharmacokinetic properties, 18 chemical compounds in FDY003 were shown to be potentially active constituents interacting with 140 BC-associated therapeutic targets to produce the pharmacological activity. Gene ontology enrichment analysis using g:Profiler indicated that the FDY003 targets were involved in the modulation of cellular processes, involving the cell proliferation, cell cycle process, and cell apoptosis. Based on a KEGG pathway enrichment analysis, we further revealed that a variety of oncogenic pathways that play key roles in the pathology of BC were significantly enriched with the therapeutic targets of FDY003; these included PI3K-Akt, MAPK, focal adhesion, FoxO, TNF, and estrogen signaling pathways. Here, we present a network-perspective of the molecular mechanisms via which herbal drugs treat BC.

Uncovering the Anti-Lung-Cancer Mechanisms of the Herbal Drug FDY2004 by Network Pharmacology.

With growing evidence on the therapeutic efficacy and safety of herbal drugs, there has been a substantial increase in their application in the lung cancer treatment. Meanwhile, their action mechanisms at the system level have not been comprehensively uncovered. To this end, we employed a network pharmacology methodology to elucidate the systematic action mechanisms of FDY2004, an anticancer herbal drug composed of Moutan Radicis Cortex, Persicae Semen, and Rhei Radix et Rhizoma, in lung cancer treatment. By evaluating the pharmacokinetic properties of the chemical compounds present in FDY2004 using herbal medicine-associated databases, we identified its 29 active chemical components interacting with 141 lung cancer-associated therapeutic targets in humans. The functional enrichment analysis of the lung cancer-related targets of FDY2004 revealed the enriched Gene Ontology terms, involving the regulation of cell proliferation and growth, cell survival and death, and oxidative stress responses. Moreover, we identified key FDY2004-targeted oncogenic and tumor-suppressive pathways associated with lung cancer, including the phosphatidylinositol 3-kinase-Akt, mitogen-activated protein kinase, tumor necrosis factor, Ras, focal adhesion, and hypoxia-inducible factor-1 signaling pathways. Overall, our study provides novel evidence and basis for research on the comprehensive anticancer mechanisms of herbal medicines in lung cancer treatment.

An Investigation of the Molecular Mechanisms Underlying the Analgesic Effect of Jakyak-Gamcho Decoction: A Network Pharmacology Study.

Herbal drugs have drawn substantial interest as effective analgesic agents; however, their therapeutic mechanisms remain to be fully understood. To address this question, we performed a network pharmacology study to explore the system-level mechanisms that underlie the analgesic activity of Jakyak-Gamcho decoction (JGd; Shaoyao-Gancao-Tang in Chinese and Shakuyaku-Kanzo-To in Japanese), an herbal prescription consisting of Paeonia lactiflora Pallas and Glycyrrhiza uralensis Fischer. Based on comprehensive information regarding the pharmacological and chemical properties of the herbal constituents of JGd, we identified 57 active chemical compounds and their 70 pain-associated targets. The JGd targets were determined to be involved in the regulation of diverse biological activities as follows: calcium- and cytokine-mediated signalings, calcium ion concentration and homeostasis, cellular behaviors of muscle and neuronal cells, inflammatory response, and response to chemical, cytokine, drug, and oxidative stress. The targets were further enriched in various pain-associated signalings, including the PI3K-Akt, estrogen, ErbB, neurotrophin, neuroactive ligand-receptor interaction, HIF-1, serotonergic synapse, JAK-STAT, and cAMP pathways. Thus, these data provide a systematic basis to understand the molecular mechanisms underlying the analgesic activity of herbal drugs.

Assessing the Anti-cancer Therapeutic Mechanism of a Herbal Combination for Breast Cancer on System-level by a Network Pharmacological Approach.

BACKGROUND/AIM: Accumulating evidence has shown therapeutic effects of herbals on breast cancer, a commonly diagnosed malignancy in women worldwide. However, their underlying mechanisms remain unclear. We aimed to explore the mode of action of a recently developed herbal combination at system-level. MATERIALS AND METHODS: We employed network pharmacological approaches to study the mechanism of a combination of three herbals, Astragalus membranaceus, Angelica gigas and Trichosanthes kirilowii by investigating active compounds and performing functional enrichment analysis for the interacting targets. RESULTS: For in silico pharmacokinetic evaluation, ten active ingredients interacted with fifty-six breast cancer-associated therapeutic targets. Functional enrichment analysis revealed that TNF, estrogen, PI3K-Akt and MAPK signaling pathways were involved in tumorigenesis and development of breast cancer. The pharmacological mechanisms might be associated with cellular effects on proliferation, cell cycle process and apoptosis. CONCLUSION: The present study provides novel insights into the system-level pharmacological mechanisms underlying a herbal combination used for breast cancer therapies.

Network Pharmacology-Based Investigation of the System-Level Molecular Mechanisms of the Hematopoietic Activity of Samul-Tang, a Traditional Korean Herbal Formula.

Hematopoiesis is a dynamic process of the continuous production of diverse blood cell types to meet the body's physiological demands and involves complex regulation of multiple cellular mechanisms in hematopoietic stem cells, including proliferation, self-renewal, differentiation, and apoptosis. Disruption of the hematopoietic system is known to cause various hematological disorders such as myelosuppression. There is growing evidence on the beneficial effects of herbal medicines on hematopoiesis; however, their mechanism of action remains unclear. In this study, we conducted a network pharmacological-based investigation of the system-level mechanisms underlying the hematopoietic activity of Samul-tang, which is an herbal formula consisting of four herbal medicines, including Angelicae Gigantis Radix, Rehmanniae Radix Preparata, Paeoniae Radix Alba, and Cnidii Rhizoma. In silico analysis of the absorption-distribution-metabolism-excretion model identified 16 active phytochemical compounds contained in Samul-tang that may target 158 genes/proteins associated with myelosuppression to exert pharmacological effects. Functional enrichment analysis suggested that the targets of Samul-tang were significantly enriched in multiple pathways closely related to the hematopoiesis and myelosuppression development, including the PI3K-Akt, MAPK, IL-17, TNF, FoxO, HIF-1, NF-kappa B, and p53 signaling pathways. Our study provides novel evidence regarding the system-level mechanisms underlying the hematopoiesis-promoting effect of herbal medicines for hematological disorder treatment.

Dual actions on gout flare and acute kidney injury along with enhanced renal transporter activities by Yokuininto, a Kampo medicine.

BACKGROUND: Prolonged hyperuricemia is associated with kidney disease or gouty arthritis. Whether Yokuininto, a commercially available Kampo medicine that has been used for osteoarthritis or rheumatoid arthritis, can exhibit anti-hyperuricemic and inflammatory effects remains elusive. In the present study, Yokuininto exerts multiple homeostatic action on serum uric acid (sUA) levels by blocking pro-inflammatory cytokine activities and inducing uricosuric function with anti-renal injury functions. METHODS: The sUA was measured in potassium oxonate (PO)-administered mice. Renal transporter uptake assays were performed using HEK293 cells overexpressing OAT1, OCT2 or OAT3, MDCKII cells overexpressing BCRP, and Xenopus oocytes overexpressing OAT3 or URAT1. Immunoblot and ELISA assays were performed to detect the molecules (OAT3, GLUT9, XO, NGAL, KIM-1 and IL-1α) in various human kidney cell lines. Cell viability analysis was performed to evaluate the cytotoxicity of Yokuininto [Ephedrine + pseudoephedrine 21.94%; Paeoniflorin 35.40% and Liquiritin 16.21% relatively measured by the ratios (HR-MS2 intensity / HR-MS1 intensity)]. RESULTS: Yokuininto (300 mg/kg) significantly reduced sUA by approximately 44% compared to that of PO-induced mice. The OAT3 levels were decreased in PO-induced hyperuricemic condition, whereas the GLUT9 transporter levels were markedly increased. However, PO did not alter the levels of URAT1. Yokuininto significantly inhibited the lipopolysaccharide (LPS)-induced secretion of IL-1α by approximately 63.2% compared to the LPS-treated macrophages. In addition, Yokuininto inhibited nitric oxide synthesis by approximately 33.7 (500 µg/mL) and 64.6% (1000 µg/mL), compared to that of LPS-treated macrophages. Yokuininto markedly increased xanthine oxidase inhibition activity. Furthermore, interleukin-1α (IL-1α), a pro-inflammatory cytokine, elevated neutrophil gelatinase-associated lipocalin (NGAL) and kidney injury molecule-1 (KIM-1) activities in LLC-PK1 cells. Expression of renal inflammatory biomarkers, NGAL and KIM-1, was reduced under the Yokuininto treatment by 36.9 and 72.1%, respectively. CONCLUSIONS: Those results suggest that Yokuininto may suppress inflammation and protect against kidney dysfunction in hyperuricemia. The present findings demonstrated that Yokuininto lowered sUA through both increased uric acid excretion and decreased uric acid production. Our results may provide a basis for the protection of prolonged hyperuricemia-associated kidney injury with uric acid-lowering agents such as Yokuininto.

Inhibitory effects of Kampo medicines, Keishibukuryogan and Shakuyakukanzoto, on the substrate uptake activities of solute carrier organic anion transporters.

The aim of this study was to assess the effects of Keishibukuryogan (K-06) and Shakuyakukanzoto (TJ-68), commercial herbal medicines, on the substrate uptake activities of renal organic anion transporters. We performed transporter uptake and cell viability assays in Xenopus oocytes and HEK293 human kidney embryonic cells treated with K-06 or TJ-68. K-06 and TJ-68 markedly inhibited the substrate uptake activities of URAT1, OAT1, and OAT3, while they did not exhibit non-cytotoxic effects. Our findings demonstrated that K-06 and TJ-68 inhibited the substrate uptake activities of renal transporters, suggesting their mechanism of action as nephroprotective agents.

Aqueous Extract of Humulus japonicus Attenuates Hyperlipidemia and Fatty Liver in Obese Mice.

In this study, the effects of Humulus japonicus (HJ) aqueous extract on 3T3-L1 preadipocytes and HepG2 cells (in vitro model) as well as on C57BL/6 mice fed on high-fat diet (HFD) (in vivo model) were evaluated. Mice fed on HFD for 12-weeks were taken the HJ water extract (HJW) at various doses, 50, 150, and 250 mg/kg, orally for 8 weeks. We have noticed the accumulation of fat globules in preadipocytes and HepG2 cells using Oil Red O staining. In addition, supplementation with HJW considerably inhibited the weight gain, lipid accumulation, and adipogenesis and decreased the size of subcutaneous adipocytes in 3T3-L1 adipocytes. Furthermore, treatment with HJW improved hyperlipidemia via decreasing the levels of serum triglyceride (TG) and low-density lipoproteins as well as the atherogenic index. Supplementation with HJW could attenuate HFD-induced lipid accumulation, increase the mRNA expressions of fatty acid synthase (FAS) and stearoyl-CoA desaturase (SCD1), and would elevate the levels of serum aspartate aminotransferase and alanine aminotransferase in mice liver. The levels of TG and FAS mRNA in HepG2 cells treated with palmitate were reduced in a dose-dependent manner. In sum, HJW could alleviate the HFD-induced obesity and decrease the dyslipidemia profiles; the keys that could contribute to cardiovascular and nonalcoholic liver diseases.

In vitro and in vivo evaluation of tissue-cultured mountain ginseng on penile erection.

BACKGROUND: Progressed tissue culture techniques have allowed us to easily obtain mass products of tissue-cultured mountain ginseng over 100 yr old (TCMG-100). We investigated the effects of TCMG-100 extract on erectile function using in vitro and in vivo studies. METHODS: To examine the relaxation effects and mechanisms of action of TCMG-100 on rabbit cavernosal strips evaluated in an organ bath. To investigate the long-term treatment effect of TCMG-100, 8-wk administration was performed. After administration of TCMG-100, intracavernosal pressure, cyclic guanosine monophosphate and nitric oxide (NO) levels of cavernosal tissue, serum testosterone level, histological observation of collagen fiber, endothelium, smooth muscle cell, and transforming growth factor-ß1 were investigated. RESULTS: TCMG-100 extract displayed dose-dependent relaxation effects on precontracted rabbit corporal smooth muscle. The TCMG-100-induced relaxation was significantly reduced by removing the endothelium, and treatment with an NO synthase inhibitor or NO scavenger. Eight weeks of TCMG-100 administration increased intracavernosal pressure in a rat model. The levels of cyclic guanosine monophosphate and NO in the corpus callosum and serum testosterone level were also increased by TCMG-100 treatment. Furthermore, histological evaluation of collagen, smooth muscle, and endothelium showed increases in endothelium and smooth muscle, and a decrease in transforming growth factor-ß1 expression. CONCLUSION: These relaxation effects on corporal smooth muscle and increased erectile function suggest that TCMG-100 might be used as an alternative herbal medicine to improve erectile function.

Effects of Aqueous Extract of Phyllostachyos Caulis in Taeniam on Longitudinal Bone Growth in Adolescent Rats.

This study examined whether treatment with Phyllostachyos Caulis in Taeniam aqueous extract improves longitudinal bone growth in adolescent male rats. Three-week-old male Sprague-Dawley rats were divided into three groups: a control group, a Phyllostachyos Caulis in Taeniam group (200 mg/kg, p.o.), and a recombinant human growth hormone group (20 µg/kg, s.c.). The total tibial length and the height of each growth plate zone were evaluated by radiography and histomorphometry. The total number of proliferative cells and 5-bromo-2'-deoxyuridine-positive cells were counted after 5-bromo-2'-deoxyuridine staining. Serum total osteocalcin levels were assayed using an enzyme-linked immunosorbent assay. The average total tibial length of the Phyllostachyos Caulis in Taeniam group was significantly longer than that of the control group. The heights of the proliferative and hypertrophic zones in the Phyllostachyos Caulis in Taeniam group were increased, and the ratio of 5-bromo-2'-deoxyuridine-positive to total cells in the proliferative zone was also increased. The serum osteocalcin, growth hormone, and insulin-like growth factor-1 levels were significantly increased in the Phyllostachyos Caulis in Taeniam group compared to the control group. Insulin-like growth factor-1 and insulin-like growth factor-1 receptor were highly expressed in the proliferative and hypertrophic zones in the Phyllostachyos Caulis in Taeniam group. The Phyllostachyos Caulis in Taeniam extract increased bone length, promoted cell proliferation, and activated the growth plate zones, which suggested that the extract could play an important role in longitudinal bone growth. Therefore, the Phyllostachyos Caulis in Taeniam extract might be a good alternative medicine to growth hormone therapy.

Beneficial Effects of Red Yeast Rice on High-Fat Diet-Induced Obesity, Hyperlipidemia, and Fatty Liver in Mice.

Obesity is a common cause of hyperlipidemia, which is a major coronary risk factor. Previous studies have shown red yeast rice (RYR) effectiveness in lowering low-density lipoprotein cholesterol. The aim of this study was to investigate the effects of RYR on obesity and hyperlipidemia. Mice were randomly separated into five groups: the control group with a normal diet, the high-fat diet (HFD) group fed a HFD without any treatment, and HFD-fed groups supplemented with RYR (1 g/kg/day for 8 weeks, 1 g/kg/day for 12 weeks, and 2.5 g/kg/day for 8 weeks). Body weight was recorded twice and food intake thrice weekly. Liver and fat pads were surgically removed and weighed. The levels of lipid parameters, liver enzymes, and leptin levels were measured. The HFD feeding resulted in obesity, which was associated with increases in body weight, liver weight, fat pad weight, liver enzymes, and plasma leptin levels with the development of hyperlipidemia. RYR prevented weight gain and fat pad weight in mice fed a HFD. RYR alleviated blood lipid parameters, liver enzymes, and leptin levels, and improved atherogenic index. These findings suggest that RYR has therapeutic potential in treating obesity and hyperlipidemia.

Clinical effect of a polysaccharide-rich extract of Acanthopanax senticosus on alcohol hangover.

The present study aimed to examine the effects polysaccharide-rich extract of Acanthopanax senticosus (PEA) on blood alcohol concentration (BAC) and hangover as well as blood lab parameters. A randomized, placebo-controlled, double-blind crossover trial was conducted. The PEA was orally administered before and after consuming alcohol 1.75 g/kg of pure alcohol. After alcohol consumption, BAC was measured for evaluation of alcohol pharmacokinetics. In the second day morning, subjects were asked to complete the Acute Hangover Scale (AHS) questionnarie. BAC results showed little difference between placebo and PEA groups, indicating that PEA does not have an effect on the pharmacokinetics of alcohol. However, several AHS items (i.e., tired, headache, dizziness, stomachache and nausea) and AHS total score were significantly improved by PEA. Blood lab parameters were significantly altered by alcohol in the placebo group. The alteration by alcohol of glucose and C-reactive protein (CRP) level was significantly attenuated by PEA. Therefore, PEA may have potential to reduce the severity of the alcohol hangover by inhibiting the alcohol-induced hypoglycemia and inflammatory response.

The role of phosphatidylcholine and deoxycholic acid in inflammation.

AIMS: Phosphatidylcholine with deoxycholic acid (PC/DA) is widely used to reduce localized fat deposits with mild adverse effects. We previously demonstrated that PC induces lipolysis with mild PMN infiltration, while DA induces adipose tissue damage. Therefore, the aim of this study was to extend our understanding of the pro-inflammatory responses of PC, DA, and PC/DA. MAIN METHODS: We evaluated the level of edema and polymononuclear (PMN) infiltration by histopathological examination. Myeloperoxidase (MPO) activity was analyzed using an MPO activity assay kit. Levels of inflammatory cytokines (IL-1ß and IL-6) and PGE2 were measured by ELISA. KEY FINDINGS: A low and high dose of PC failed to induce an inflammatory response, whereas DA led to an intense inflammatory response in a dose dependent manner. Combined PC/DA treatment resulted in a mild inflammatory response that was notably less severe than higher DA. Together, these results demonstrated that DA plays a role in inflammation caused by combined PC/DA. Histopathological examination and measurement of MPO activity indicated that DA was the primary cause of edema and PMN infiltration. Further, increased levels of cytokines (IL-1ß and IL-6) and PGE2 demonstrated that DA might directly induce inflammation, whereas PC alone has no effect on inflammation. SIGNIFICANCE: These results indicate that DA rather than PC is responsible for inflammation, and that PC may not aggravate inflammatory responses induced by DA. Thus, the results of this study suggest that the adverse effects of PC/DA during localized fat treatment may be solely due to DA.

Evidence for the Primo Vascular System above the Epicardia of Rat Hearts.

We for the first time reported evidence for the existence of a novel network, a PVS, abovethe epicardium of the rat heart. (1) We were consecutively able to visualize the PVs and the PNs above the epicardial spaces of five rats' hearts by using Cr-Hx spraying or injection. (2) Hematoxylin and eosin (H&E) and toluidine blue staining of the PVs and the PNs showed that they consisted of a basophilic matrix; specifically the PNs contained several mast cells, some of which were degranulating into pericardial space. Also, 4', 6-diamidino-2 phenylindole (DAPI) images of the PVs and the PNs showed that they contained various kinds of cells. (3) Transmission electron microscopic (TEM) longitudinal image of the PVs showed that the sinuses contained many granules with high-electron-density cores in parallel with putative endothelial cells. (4) TEM images of the PNs demonstrated that they consisted of lumen-containing cells surrounded by fibers and that they had mast cells that were degranulating toward the epicardium of the rat heart. The above data suggest that mast-cells-containing novel network exists above the epicardium of the rat heart.

Inhibitory effects of polysaccharide-rich extract of Phragmites rhizoma on atopic dermatitis-like skin lesions in NC/Nga mice.

AIMS: Phragmites rhizoma was reported to have anti-oxidative and free radical scavenging activity. It also has been traditionally used to suppress inflammation. In the present study, we aimed to evaluate the topical effects of the polysaccharide-rich extract of P. rhizoma (PEP) on atopic dermatitis. MAIN METHODS: We induced AD-like skin lesions by an extract of the house-dust mite Dermatophagoides farinae (Dfb) in NC/Nga mice, and then performed macroscopic analysis, immunohistochemical staining and measurement of total serum IgE and cytokine production by ELISA. KEY FINDINGS: Topically applied PEP suppressed dermatitis with a decrease in dermatitis score and scratch number. The histological manifestations of atopic skin lesions including thickened epidermis and increased numbers of mast cells, polymorphonuclear leukocytes and nerve fibers were significantly attenuated. The activation of IgE and the levels of cytokines such as IFN-γ IL-4 and IL-10 were also decreased. SIGNIFICANCE: Our results indicated that PEP might have an inhibitory effect on atopic dermatitis-like lesion and be a promising natural resource in the treatment of atopic dermatitis.

Visualization of the network of primo vessels and primo nodes above the pia mater of the brain and spine of rats by using Alcian blue.

By spraying and injecting Alcian blue into the lateral ventricle, we were able to visualize the network of the nerve primo vascular system above the pia mater of the brain and spine of rats. Staining these novel structures above the pia mater with 4',6-diamidino-2-phenylindole demonstrated that they coexisted in cellular and extracellular DNA forms. The cellular primo node consisted of many cells surrounded by rod-shaped nuclei while the extracellular primo node had a different morphology from that of a general cell in terms of DNA signals, showing granular DNA in a threadlike network of extracellular DNA. Also, differently from F-actin in general cells, the F-actin in the primo vessel was short and rod-shaped. Light and transmission electron microscopic images of the PN showed that the nerve primo vascular system above the pia mater of the brain and spine was a novel dynamic network, suggesting the coexistence of DNA and extracellular DNA. Based on these data, we suggest that a novel dynamic system with a certain function exists above the pia mater of the central nerve system. We also discuss the potential of this novel network system in the brain and spine as related to acupuncture meridians and neural regeneration.

Growth of microgranules into cell-like structures in fertilized chicken eggs: hypothesis for a mitosis-free alternative pathway.

According to Bonghan Kim's theory of anatomical reality for acupuncture meridians, DNA microgranules known as Sanals are key functional components in the primo vascular system (formerly the Bonghan system). To investigate this issue, we developed a new system, an incubator bound to a phase-contrast microscope, in which we cultivated and then observed for 10 hours microgranules taken from 3-day-old chick embryos and from blastoderms of fertilized chicken eggs. With this system, we found that, over time, the microgranules grew in circular patterns to become cell-like structures. In the embryo specimens, we found two distinctive microgranule growths, which developed into cell-like structures over 10 hours. In the first case, a microgranule of about 1.0 µm in size developed into a 3.3-µm-sized cell-like structure, with a pattern of concentric circles. The growth rate of the diameter of the first microgranule was, on average, 0.23 µm/hour. In the second case, a 2.5-µm-sized microgranule developed into a 5.4-µm-sized cell-like structure, which also exhibited a pattern of concentric circles. The average growth rate of the diameter of the second microgranule was 0.31 µm/hour. In the blastoderm specimens from the fertilized chicken egg, we also found three distinctive concentric growths. Interestingly, one of the three blastoderm microgranules grew very quickly, from about 2.5 µm in size to about 5.5 µm in size during 5 minutes of incubation. This was followed by steady growth to about 7.0 µm in size during the next 10 hours of incubation. In the final step of our investigation, we confirmed that the cell-like structures that had grown from the microgranules stained by acridine orange had DNA signals. We believe that the data obtained with our experimental method provide a clue that a mitosis-free alternative pathway for cell formation may, indeed, exist. We also suggest that this new function of microgranules (Sanals) might be related with the acupuncture meridian called the primo vascular system.