Effect of Oral Skim Milk Administration on Skeletal Muscle Protein Synthesis after Total Gastrectomy in Rat.

Leucine is a branched-chain amino acid that is present in protein, and it is an essential factor in activating the mechanistic target of the rapamycin complex 1 signaling pathway and increasing muscle protein synthesis. However, the loss of digestive function after total gastrectomy leads to impaired protein absorption, potentially failing to stimulate muscle protein synthesis. Therefore, this study aimed to investigate whether muscle protein synthesis is enhanced by oral skim milk administration after total gastrectomy. Male Sprague Dawley rats were divided into total gastrectomy (TG) and sham surgery (S) groups. After five weeks postoperatively, we orally administered skim milk to achieve 3.1 g protein/kg body weight and collected blood and gastrocnemius muscle. The gastrocnemius muscle weight was significantly lower in the TG group than in the S group (p < 0.05). The increase in plasma leucine concentration was significantly lower in the TG group than in the S group (p < 0.05). The skeletal muscle protein synthesis and the phosphorylation of p70S6K and 4E-BP1 showed a similar increase in both groups. Even after TG, muscle protein synthesis was stimulated by consuming skim milk, accompanied by a sufficient rise in plasma leucine concentration.

Association of fat-to-muscle mass ratio with physical activity and dietary protein, carbohydrate, sodium, and fiber intake in a cross-sectional study.

Higher fat-to-muscle mass ratio (FMR) is reported to be a risk factor for various diseases, including type 2 diabetes and cardiovascular diseases, and mortality. Although this association suggests that reducing FMR may help to prevent certain diseases and mortality, the relationship between FMR and lifestyle factors is unclear. Therefore, we performed a cross-sectional study with the aim to elucidate this relationship. This cross-sectional study included 1518 healthy Japanese adults aged 30 to 64 years. We measured FMR in the whole body, arms, legs, and trunk and assessed various lifestyle factors. Then, we performed forced entry multiple regression analyses for FMR with the following variables: sex, age, physical activity, dietary intake, sleep quality, cigarette smoking, stress levels, and body mass index. As a result, whole-body and regional FMRs were correlated with female sex (ß = 0.71); age (ß = 0.06); physical activity (ß = - 0.07); dietary intake of protein (ß = - 0.12), carbohydrate (ß = 0.04), sodium (ß = 0.13), and fiber (ß = - 0.16); and body mass index (ß = 0.70). The results suggest that in the Japanese middle-aged population, low FMR is associated with certain lifestyle factors, i.e. higher physical activity and a diet with higher protein and fiber and lower carbohydrate and sodium, independent of age, sex, and body mass index.

The effects of resistance exercise and leucine-enriched essential amino acid supplementation on muscle mass and physical function in post-gastrectomy patients: a pilot randomized controlled trial.

[Purpose] In gastric cancer patients, low muscle mass decreases overall survival and quality of life (QOL). Resistance exercise with leucine-enriched essential amino acid (LEAA) supplementation may prevent muscle mass loss. This study was aimed at determining whether resistance exercise with LEAA supplementation prevents muscle mass loss in post-gastrectomy patients. [Participant and Methods] We conducted a single-center, open-label, randomized controlled pilot trial. Ten participants who underwent gastrectomy were divided into two groups. The intervention group underwent resistance exercise at 70% of one repetition maximum and received a supplement of 3 g of LEAA twice daily for 15 days, while the control group received standard care. We compared changes in muscle mass, physical function (muscle strength and continuous walking distance), and QOL between the groups. [Results] We found good adherence and participation rates in both groups. We failed to detect a significant difference in muscle mass between the groups. The intervention group showed significant improvements in muscle strength and QOL, while the control group showed no significant changes. [Conclusion] We failed to detect a significant difference in muscle mass due to resistance exercise with LEAA supplementation in post-gastrectomy patients. However, resistance exercise with LEAA supplementation might be beneficial for muscle strength recovery and QOL improvements.

Intramuscular injection of mesenchymal stem cells augments basal muscle protein synthesis after bouts of resistance exercise in male mice.

Skeletal muscle mass is critical for activities of daily living. Resistance training maintains or increases muscle mass, and various strategies maximize the training adaptation. Mesenchymal stem cells (MSCs) are multipotent cells with differential potency in skeletal muscle cells and the capacity to secrete growth factors. However, little is known regarding the effect of intramuscular injection of MSCs on basal muscle protein synthesis and catabolic systems after resistance training. Here, we measured changes in basal muscle protein synthesis, the ubiquitin-proteasome system, and autophagy-lysosome system-related factors after bouts of resistance exercise by intramuscular injection of MSCs. Mice performed three bouts of resistance exercise (each consisting of 50 maximal isometric contractions elicited by electrical stimulation) on the right gastrocnemius muscle every 48 h, and immediately after the first bout, mice were intramuscularly injected with either MSCs (2.0 × 106 cells) labeled with green fluorescence protein (GFP) or vehicle only placebo. Seventy-two hours after the third exercise bout, GFP was detected only in the muscle injected with MSCs with concomitant elevation of muscle protein synthesis. The injection of MSCs also increased protein ubiquitination. These results suggest that the intramuscular injection of MSCs augmented muscle protein turnover at the basal state after consecutive resistance exercise.

Comparative Study of Postmortem MRI and Pathological Findings in Malignant Brain Tumors.

This study compared magnetic resonance imaging (MRI) findings of postmortem brain specimens with neuropathological findings to evaluate the value of postmortem MRI. Postmortem MRI was performed on five formalin-fixed whole brains with malignant tumors. Postmortem T2-weighted images detected all neuropathological abnormalities as high-signal regions but also showed histological tumor invasion in areas without edema. Tumor lesions with high necrosis and edema showed high signal intensity on T2-weighted images; in three cases, lesion enlargement was detected on the final prenatal imaging and postmortem MRI. Disease progression immediately before death may have contributed to this difference. In conclusion, the correlation between MRI and neuropathological findings facilitates understanding of the mechanisms responsible for MRI abnormalities. Increased free water due to edema, necrosis, and brain tissue injury can explain the increased signal intensity observed on T2-weighted images. Postmortem MRI may contribute to effective pathology by identifying subtle abnormalities prior to brain dissection.

Evaluation of Treatment Strategies for Male Prolactin-Secreting Pituitary Neuroendocrine Tumors.

Prolactin-secreting pituitary neuroendocrine tumors (PitNETs) are more common in women. Male patients may also have few symptoms and have macroadenomas extending outside the sella turcica. This study aimed to report the results of cabergoline treatment in male patients with prolactin-secreting PitNET. The study included nine male patients aged 26-65 years (median, 46 years) diagnosed with prolactin-secreting PitNETs. The age at onset, prolactin values, tumor size, symptoms, and treatment were assessed. The mean prolactin value at the initial presentation was 2734.6 ng/mL, and the mean maximum tumor diameter was 40.4 mm. Visual field disturbance was the most common symptom (44.4%), followed by headaches (33.3%), asymptomatic symptoms (11.1%), and galactorrhea (11.1%). Eight patients responded to cabergoline treatment with normalization of prolactin levels and tumor shrinkage. One patient did not respond to the cabergoline treatment and required surgical intervention. There were no cases of cerebrospinal fluid leakage. Cabergoline was found to be an effective treatment for male prolactin-secreting PitNETs.

Controlled Release of Oxygen from Calcium Peroxide in a Weak Acidic Condition by Stabilized Amorphous Calcium Carbonate Nanocoating for Biomedical Applications.

Calcium peroxide (CaO2) has recently attracted much attention as an oxygen-releasing biomaterial for tissue engineering. CaO2 has also been used in cancer therapies, such as photodynamic therapy. However, the uncontrollability of oxygen release after immersion in water is a challenge. Furthermore, the nanoscale surface chemistry of CaO2 on the oxygen release properties under cell culture conditions has not been taken into account in these applications. Herein, we report the stabilized amorphous calcium carbonate (ACC) nanocoating on CaO2 in a cell culture medium, which suppressed the reaction between CaO2 and water. Stabilized ACC was produced by the reaction between calcium hydroxide (Ca(OH)2) derived from CaO2 and sodium hydrogen carbonate (NaHCO3) including sodium dihydrogen phosphate (NaH2PO4) in a cell culture medium. In contrast, surface modification of CaO2 by calcium carbonate crystals was difficult due to the crystallization process via dissolution-reprecipitation. Strikingly, ACC-CaO2 showed pH-dependent oxygen release in a cell culture medium probably because of the dissolution of ACC under weak acidic condition. Since the environments in ischemic tissue and cancer are weakly acidic, our findings should be important for understanding and designing properties related to biomaterials and drugs using CaO2.

Cooling-promoted myogenic differentiation of murine bone marrow mesenchymal stem cells through TRPM8 activation in vitro.

TRPM8 agonist has been reported to promote osteogenic differentiation of mesenchymal stem cells (MSCs), therefore we evaluated whether cooling-induced activation of TRPM8 promotes myogenic differentiation of MSCs. We used 5-azacytidine as a myogenic differentiation inducer in murine bone marrow-derived MSCs. Addition of menthol, a TRPM8 agonist, to the differentiation induction medium significantly, increased the percentage of MyoD-positive cells, a specific marker of myogenic differentiation. We performed intracellular Ca2+ imaging experiments using fura-2 to confirm TRPM8 activation by cooling stimulation. The results confirmed that intracellular Ca2+ concentration ([Ca2+ ]i) increases due to TRPM8 activation, and TRPM8 antagonist inhibits increase in [Ca2+ ]i at medium temperatures below 19°C. We also examined the effect of cooling exposure time on myogenic differentiation of MSCs using an external cooling stimulus set at 17°C. The results showed that 60 min of cooling had an acceleratory effect on differentiation (2.18 ± 0.27 times). We observed that the TRPM8 antagonist counteracted the differentiation-promoting effect of the cooling. These results suggest that TRPM8 might modulate the multiple differentiation pathways of MSCs, and that cooling is an effective way of activating TRPM8, which regulates MSCs differentiation in vitro.

Biventricular repair using subvalvular techniques for unbalanced atrioventricular septal defect.

We report the case of a 19-month-old girl with a right-dominant unbalanced atrioventricular septal defect and severe right-sided atrioventricular valve regurgitation who underwent biventricular repair using basal chordae resection, artificial chordae reconstruction and a left-sided atrioventricular valvuloplasty. At 14-month postoperative follow-up, the patient had minimal heart failure, gained weight and adapted to biventricular circulation.

Efficacy and Safety of 6-Month High Dietary Protein Intake in Hospitalized Adults Aged 75 or Older at Nutritional Risk: An Exploratory, Randomized, Controlled Study.

The aim of this study was to investigate the effects of increased dietary protein in daily-life settings in Japan for 6 months on the activities of daily living (ADL) in adults aged 75 or older at nutritional risk. The study was an open-label, exploratory, randomized controlled trial conducted at seven hospitals in Japan. The study participants were adults aged 75 or older who were hospitalized for treatable cancer, pneumonia, fractures, and/or urinary-tract infection at nutritional risk. The primary outcome was change in grip strength, skeletal muscle, and ADL indices (Barthel index, Lawton score). One hundred sixty-nine patients were randomly assigned to the intensive care (IC) or standard care (SC) group; the protein intake goals (g/kgw/day) were 1.5 for IC and 1.0 for SC. There was a significant improvement in grip strength only in the IC group (1.1 kg: 95% CI 0.1 to 2.1) (p = 0.02). While the skeletal muscle index and ADL indices were not significantly improved in either group, the improvement ratio tended to be greater in the IC group. There was no decrease in renal function in either group. Thus, intervention of increased dietary protein in daily-life settings for 6 months in adults aged 75 or older with treatable cancer, pneumonia, fractures, and/or urinary-tract infection and at nutritional risk may be effective in ameliorating loss of muscle strength.

Necrostatin-1 Attenuates Delayed Paraplegia after Transient Spinal Cord Ischemia in Rabbits by Inhibiting the Upregulation of Receptor-Interacting Protein Kinase 1 and 3.

BACKGROUND: Delayed-onset paraplegia is a disastrous complication after thoracoabdominal aortic open surgery and thoracic endovascular aortic repair. Studies have revealed that transient spinal cord ischemia caused by temporary occlusion of the aorta induces delayed motor neuron death owing to apoptosis and necroptosis. Recently, necrostatin-1 (Nec-1), a necroptosis inhibitor, has been reported to reduce cerebral and myocardial infarction in rats or pigs. In this study, we investigated the efficacy of Nec-1 in delayed paraplegia after transient spinal cord ischemia in rabbits and assessed the expression of necroptosis- and apoptosis-related proteins in motor neurons. METHODS: This study used rabbit transient spinal cord ischemia models using a balloon catheter. They were divided into a vehicle-treated group (n = 24), Nec-1-treated group (n = 24), and sham-controls (n = 6). In the Nec-1-treated group, 1 mg/kg of Nec-1 was intravascularly administered immediately before ischemia induction. Neurological function was assessed using the modified Tarlov score, and the spinal cord was removed 8 hr and 1, 2, and 7 days after reperfusion. Morphological changes were examined using hematoxylin and eosin staining. The expression levels of necroptosis-related proteins (receptor-interacting protein kinase [RIP] 1 and 3) and apoptosis-related proteins (Bax and caspase-8) were assessed using western blotting and histochemical analysis. We also performed double-fluorescence immunohistochemical studies of RIP1, RIP3, Bax, and caspase-8. RESULTS: Neurological function significantly improved in the Nec-1-treated group compared with that in the vehicle-treated group 7 days after reperfusion (median 3 and 0, P = 0.025). Motor neurons observed 7 days after reperfusion were significantly decreased in both groups compared with the sham group (vehicle-treated, P < 0.001; Nec-1-treated, P < 0.001). However, significantly more motor neurons survived in the Nec-1-treated group than in the vehicle-treated group (P < 0.001). Western blot analysis revealed RIP1, RIP3, Bax, and caspase-8 upregulation 8 hr after reperfusion in the vehicle-treated group (RIP1, P = 0.001; RIP3, P = 0.045; Bax, P = 0.042; caspase-8, P = 0.047). In the Nec-1-treated group, the upregulation of RIP1 and RIP3 was not observed at any time point, whereas that of Bax and caspase-8 was observed 8 hr after reperfusion (Bax, P = 0.029; caspase-8, P = 0.021). Immunohistochemical study revealed the immunoreactivity of these proteins in motor neurons. Double-fluorescence immunohistochemistry revealed the induction of RIP1 and RIP3, and that of Bax and caspase-8, in the same motor neurons. CONCLUSIONS: These data suggest that Nec-1 reduces delayed motor neuron death and attenuates delayed paraplegia after transient spinal cord ischemia in rabbits by selectively inhibiting necroptosis of motor neurons with minimal effect on their apoptosis.

Single-Cell Analysis of Unidirectional Migration of Glioblastoma Cells Using a Fiber-Based Scaffold.

Glioblastoma (GBM) is a malignant incurable brain tumor in which immature neoplastic cells infiltrate brain tissue by spreading along nerve fibers. The aim of the study was to compare the migration abilities of glioma cells with those of other cancer cells and elucidate the migratory profiles underlying the differential migration of glioma cells using a fiber-based quantitative migration assay. Here, wound healing and transwell assays were used to assess cell mobility in four cell lines: U87-MG glioblastoma cells, MDA-MB-231 breast cancer cells, HCT116 colorectal cancer cells, and MKN45 gastric cancer cells. We also assessed cell mobility using a fiber model that mimics nerve fibers. Time-lapse video microscopy was used to observe cell migration and morphology. The cytoskeleton arrangement was assessed in the fiber model and compared with that in the conventional cell culture model. The conventional evaluation of cell migration ability revealed that the migration ability of breast cancer and glioblastoma cell lines was higher than that of colon cancer and gastric cancer cell lines. The fiber model confirmed that the glioblastoma cell line had a significantly higher migration ability than other cell lines. Tubulin levels were significantly higher in the glioblastoma cells than in other cell lines. In conclusion, the developed fiber-based culture model revealed the specific migratory profile of GBM cells during invasion.

Associated factors and effects of comorbid atrial fibrillation in hypertensive patients due to primary aldosteronism.

The incidence of atrial fibrillation (AF) and risk of cardiovascular events are reportedly higher in patients with primary aldosteronism (PA) than essential hypertension. However, associated factors of comorbid AF and cardiovascular events in PA patients after PA treatment remain unclear. This nationwide registration study included PA patients ≥20 years old. Incident cardiovascular events were observed with a mean follow-up of approximately 3 years. A total of 3654 patients with PA were included at the time of analysis. Prevalence of AF was 2.4%. PA patients with AF were older, more frequently male and had longer duration of hypertension than those without AF. No significant difference in basal plasma and adrenal venous aldosterone concentration, renin activity, potassium concentration, confirmatory tests of PA, laterality or surgery rate were seen between groups. Logistic regression analysis showed age, male sex, cardiothoracic ratio, past history of coronary artery disease and heart failure were independent factors associated with AF. PA patients with AF showed a higher frequency of cardiovascular events than those without AF (P < 0.001). Multivariate Cox analyses demonstrated AF in addition to older age, duration of hypertension, body mass index and chronic kidney disease as independent prognostic factors for cardiovascular events after PA treatment. Incidence of cardiovascular events were significantly lower in PA patients with AF than AF patients from the Fushimi registry during follow-up after adjusting age, sex and systolic blood pressure. Early diagnosis of PA may prevent AF and other cardiovascular events in PA patients by shortening the duration of hypertension and appropriate PA treatment.

A case of toxic epidermal necrolysis comorbid with severe burns.

Toxic epidermal necrolysis (TEN) and severe burns both have high mortality rates, but coexistence is extremely rare. The specificity of developing TEN in burn patients is not well understood and its treatment strategy is not established. Case Presentation: A 68-year-old man was carried to our hospital with severe burns covering 35% of his body surface area. He developed bacteremia during treatment of burns and required antimicrobial therapy. However, erythema appeared on the trunk and upper limbs and rapidly spread to the extremities, leading to a diagnosis of TEN. The rash gradually improved after terminating antimicrobial therapy and administrating of 1,000 mg/day methylprednisolone for 3 days. The rash caused by TEN was confined to non-burned areas, suggesting that TEN may less likely occur at burn sites. Conclusion: It is necessary to pay attention because burn patients can develop TEN concomitantly. Corticosteroids therapy may be effective for TEN even in severe burn patients.

Quantitative Analysis of Collective Migration by Single-Cell Tracking Aimed at Understanding Cancer Metastasis.

Metastasis is a major complication of cancer treatments. Studies of the migratory behavior of cells are needed to investigate and control metastasis. Metastasis is based on the epithelial-mesenchymal transition, in which epithelial cells acquire mesenchymal properties and the ability to leave the population to invade other regions of the body. In collective migration, highly migratory "leader" cells are found at the front of the cell population, as well as cells that "follow" these leader cells. However, the interactions between these cells are not well understood. We examined the migration properties of leader-follower cells during collective migration at the single-cell level. Different mixed ratios of "leader" and "follower" cell populations were compared. Collective migration was quantitatively analyzed from two perspectives: cell migration within the colony and migration of the entire colony. Analysis of the effect of the cell mixing ratio on migration behavior showed that a small number of highly migratory cells enhanced some of the migratory properties of other cells. The results provide useful insights into the cellular interactions in collective cell migration of cancer cell invasion.

Time-Series Clustering of Single-Cell Trajectories in Collective Cell Migration.

Collective invasion drives multicellular cancer cells to spread to surrounding normal tissues. To fully comprehend metastasis, the methodology of analysis of individual cell migration in tissue should be well developed. Extracting and classifying cells with similar migratory characteristics in a colony would facilitate an understanding of complex cell migration patterns. Here, we used electrospun fibers as the extracellular matrix for the in vitro modeling of collective cell migration, clustering of mesenchymal and epithelial cells based on trajectories, and analysis of collective migration patterns based on trajectory similarity. We normalized the trajectories to eliminate the effect of cell location on clustering and used uniform manifold approximation and projection to perform dimensionality reduction on the time-series data before clustering. When the clustering results were superimposed on the trajectories before normalization, the results still exhibited positional similarity, thereby demonstrating that this method can identify cells with similar migration patterns. The same cluster contained both mesenchymal and epithelial cells, and this result was related to cell location and cell division. These data highlight the reliability of this method in identifying consistent migration patterns during collective cell migration. This provides new insights into the epithelial-mesenchymal interactions that affect migration patterns.

Development of non-adherent cell-enclosing domes with enzymatically cross-linked hydrogel shell.

Non-adherent cells, such as hematopoietic cells and lymphocytes, are important research subjects in medical and biological fields. Therefore, a system that enables the handling of non-adherent cells in solutions in the same manner as that of adhering cells during medium exchange, exposure to chemicals, washing, and staining in imaging applications would be useful. Here, we report a 'Cell Dome' platform in which non-adherent cells can be enclosed and grown in the cavities of about 1 mm diameter and 270µm height. The domes consist of an alginate-based hydrogel shell of 90µm thickness. Cell Domes were formed on glass plates by horseradish peroxidase-mediated cross-linking. Human leukaemia cell line K562 cells enclosed in Cell Domes were stable for 29 days with every 2-3 days of medium change. The enclosed cells grew in the cavities and were stained and differentiated with reagents supplied from the surrounding medium. Additionally, K562 cells that filled the cavities (a 3D microenvironment) were more hypoxic and highly resistant to mitomycin C than those cultured in 2D. These findings demonstrate that the 'Cell Dome' may be a promising tool for conveniently culturing and evaluating non-adherent cells.

The Efficacy and Complications of Preoperative Embolization of Metastatic Spinal Tumors: Risk of Paralysis after Embolization.

Introduction: This study investigated the efficacy and complications of preoperative embolization for spinal metastatic tumors, focusing on the etiology of post-embolization paralysis. Methods: We retrospectively reviewed the data of 44 consecutive patients with spinal metastases treated between September 2012 and December 2020. Intraoperative blood loss and postoperative transfusion requirement were compared between the embolization (+) and (-) groups. Complications associated with embolization were reviewed. Results: Overall, 30 patients (68%) underwent preoperative embolization. All the patients in both groups underwent palliative posterior decompression and fusion. The mean intraoperative blood loss in the overall population was 359 ml (range, minimum-2190 ml) and was 401 ml and 267 ml in the embolization (+) and embolization (-) groups, respectively. Four patients (9%) (2 patients from each group) required blood transfusion. There were no significant between-group differences in blood loss and blood transfusion requirements. All 7 patients with hypervascular tumors were in the embolization (+) group. Two patients experienced muscle weakness in the lower extremities on days 1 and 3 after embolization. There were metastases in T5 and T1-2, and magnetic resonance imaging after embolization showed slight exacerbation of spinal cord compression. The patients showed partial recovery after surgery. Conclusions: With the predominance of hypervascular tumors in the embolization (+) group, preoperative embolization may positively affect intraoperative bleeding. Embolization of metastatic spinal tumors may pose a risk of paralysis. Although the cause of paralysis remains unclear, it might be due to the aggravation of spinal cord compression. Considering this risk of paralysis, we advocate performing surgery as soon as possible after embolization.

A peptide-mediated, multilateral molecular dialogue for the coordination of pollen wall formation.

The surface of pollen grains is reinforced by pollen wall components produced noncell autonomously by tapetum cells that surround developing pollen within the male floral organ, the anther. Here, we show that tapetum activity is regulated by the GASSHO (GSO) receptor-like kinase pathway, controlled by two sulfated peptides, CASPARIAN STRIP INTEGRITY FACTOR 3 (CIF3) and CIF4, the precursors of which are expressed in the tapetum itself. Coordination of tapetum activity with pollen grain development depends on the action of subtilases, including AtSBT5.4, which are produced stage specifically by developing pollen grains. Tapetum-derived CIF precursors are processed by subtilases, triggering GSO-dependent tapetum activation. We show that the GSO receptors act from the middle layer, a tissue surrounding the tapetum and developing pollen. Three concentrically organized cell types, therefore, cooperate to coordinate pollen wall deposition through a multilateral molecular dialogue.

Cell Dome as an Evaluation Platform for Organized HepG2 Cells.

Human-hepatoblastoma-derived cell line, HepG2, has been widely used in liver and liver cancer studies. HepG2 spheroids produced in a three-dimensional (3D) culture system provide a better biological model than cells cultured in a two-dimensional (2D) culture system. Since cells at the center of spheroids exhibit specific behaviors attributed to hypoxic conditions, a 3D cell culture system that allows the observation of such cells using conventional optical or fluorescence microscopes would be useful. In this study, HepG2 cells were cultured in "Cell Dome", a micro-dome in which cells are enclosed in a cavity consisting of a hemispherical hydrogel shell. HepG2 cells formed hemispherical cell aggregates which filled the cavity of Cell Domes on 18 days of culture and the cells could continue to be cultured for 29 days. The cells at the center of hemispherical cell aggregates were observed using a fluorescence microscope. The cells grew in Cell Domes for 18 days exhibited higher Pi-class Glutathione S-Transferase enzymatic activity, hypoxia inducible factor-1α gene expression, and higher tolerance to mitomycin C than those cultured in 2D on tissue culture dishes (* p < 0.05). These results indicate that the center of the glass adhesive surface of hemispherical cell aggregates which is expected to have the similar environment as the center of the spheroids can be directly observed through glass plates. In conclusion, Cell Dome would be useful as an evaluation platform for organized HepG2 cells.