[Monitoring for Spine and Spinal Cord Surgery].

Injuries of the spine and nerve root may occur during spine or spinal cord surgery, which can induce serious neurological deficits. Intraoperative monitoring plays important roles in monitoring the nerve function in various surgical manipulations, such as surgical positioning, mechanical compression, and tumor removal. This monitoring issues a warning to neuronal injuries at an early stage, and surgeons are able to prevent postoperative complications. The appropriate monitoring systems should be chosen with compatibility to the disease, surgical procedure, and lesion localization. The team should share the significance of monitoring and understand the timing of stimulation to perform a safe surgery. In this paper, we overview various intraoperative monitoring techniques and pitfalls in spine and spinal cord surgeries based on cases from our hospital.

Simple propagation method for resident macrophages by co-culture and subculture, and their isolation from various organs.

BACKGROUND: Resident macrophages (Mø) originating from yolk sac Mø and/or foetal monocytes colonise tissues/organs during embryonic development. They persist into adulthood by self-renewal at a steady state, independent of adult monocyte inputs, except for those in the intestines and dermis. Thus, many resident Mø can be propagated in vitro under optimal conditions; however, there are no specific in vitro culture methods available for the propagation of resident Mø from diverse tissues/organs. RESULTS: We provided a simple method for propagating resident Mø derived from the liver, spleen, lung, and brain of ICR male mice by co-culture and subculture along with the propagation of other stromal cells of the respective organs in standard culture media and successfully demonstrated the propagation of resident Mø colonising these organs. We also proposed a simple method for segregating Mø from stromal cells according to their adhesive property on bacteriological Petri dishes, which enabled the collection of more than 97.6% of the resident Mø from each organ. Expression analyses of conventional Mø markers by flow cytometry showed similar expression patterns among the Mø collected from the organs. CONCLUSION: This is the first study to clearly provide a practical Mø propagation method applicable to resident Mø of diverse tissues and organs. Thus, this novel practical Mø propagation method can offer broad applications for the use of resident Mø of diverse tissues and organs.

Impairment of endothelial-mesenchymal transformation during atrioventricular cushion formation in Tmem100 null embryos.

BACKGROUND: Endothelial-mesenchymal transformation (EndMT) is essential for endocardial cushion formation during cardiac morphogenesis. We recently identified Tmem100 as an endothelial gene indispensable for vascular development. In this study, we further investigated its roles for EndMT during atrioventricular canal (AVC) cushion formation. RESULTS: Tmem100 was expressed in AVC endocardial cells, and Tmem100 null embryos showed severe EndMT defect in the AVC cushions. While calcineurin-dependent suppression of vascular endothelial growth factor (VEGF) expression in the AVC myocardium is important for EndMT, significant up-regulation of Vegfa expression was observed in Tmem100 null heart. EndMT impaired in Tmem100 null AVC explants was partially but significantly restored by the expression of constitutively-active calcineurin A, suggesting dysregulation of myocardial calcineurin-VEGF signaling in Tmem100 null heart. Moreover, Tmem100 null endocardial cells in explant culture did not show EndMT in response to the treatment with myocardium-derived growth factors, transforming growth factor ß2 and bone morphogenetic protein 2, indicating involvement of an additional endocardial-specific abnormality in the mechanism of EndMT defect. The lack of NFATc1 nuclear translocation in endocardial cells of Tmem100 null embryos suggests impairment of endocardial calcium signaling. CONCLUSIONS: The Tmem100 deficiency causes EndMT defect during AVC cushion formation possibly via disturbance of multiple calcium-related signaling events.

An important role of endothelial hairy-related transcription factors in mouse vascular development.

The Hairy-related transcription factor family of Notch- and ALK1-downstream transcriptional repressors, called Hrt/Hey/Hesr/Chf/Herp/Gridlock, has complementary and indispensable functions for vascular development. While mouse embryos null for either Hrt1/Hey1 or Hrt2/Hey2 did not show early vascular phenotypes, Hrt1/Hey1; Hrt2/Hey2 double null mice (H1(ko) /H2(ko) ) showed embryonic lethality with severe impairment of vascular morphogenesis. It remained unclear, however, whether Hrt/Hey functions are required in endothelial cells or vascular smooth muscle cells. In this study, we demonstrate that mice with endothelial-specific deletion of Hrt2/Hey2 combined with global Hrt1/Hey1 deletion (H1(ko) /H2(eko) ) show abnormal vascular morphogenesis and embryonic lethality. Their defects were characterized by the failure of vascular network formation in the yolk sac, abnormalities of embryonic vascular structures and impaired smooth muscle cell recruitment, and were virtually identical to the H1(ko) /H2(ko) phenotypes. Among signaling molecules implicated in vascular development, Robo4 expression was significantly increased and activation of Src family kinases was suppressed in endothelial cells of H1(ko) /H2(eko) embryos. The present study indicates an important role of Hrt1/Hey1 and Hrt2/Hey2 in endothelial cells during early vascular development, and further suggests involvement of Robo4 and Src family kinases in the mechanisms of embryonic vascular defects caused by the Hrt/Hey deficiency.

Concomitant Administration of Vancomycin with a High Dose of Meropenem May Cause Acute Kidney Injury.

Coadministering two different classes of antibiotics as empirical therapy can be critical in treating healthcare-associated infections in hospitals. Herein, we report a case of acute kidney injury (AKI) caused by coadministration of vancomycin with high-dose meropenem that manifested as a rapid increase in serum creatinine levels and an associated increase in vancomycin trough concentrations. The patient was diagnosed with meningioma at 50 years and was followed up regularly. The patient underwent surgery and antibiotic treatment between 63 and 66 years for suspected meningitis and pneumonia. Coadministration of vancomycin with high-dose meropenem (6.0 g/day) caused AKI; however, no AKI occurred when vancomycin was administered alone or with a low dose of meropenem (1.5 or 3.0 g/day). To our knowledge, this report is the first to show that administering different dosages of meropenem in combination with vancomycin may contribute to the risk of developing AKI. We suggest that coadministered vancomycin and high-dose meropenem (6.0 g/day) may increase the risk of AKI. Our report adds to the limited literature documenting the coadministration of vancomycin with varying doses of meropenem and its impact on the risk of AKI and highlights the importance of investigating AKI risk in response to varying dosages of meropenem when it is coadministered with vancomycin.

Clinical and radiographic characteristics of patients with non-functioning pituitary adenomas categorized according to their serum prolactin concentration: novel predictors of postoperative transient diabetes insipidus following surgery.

PURPOSE: Non-functioning pituitary adenomas (NFPAs) are often associated with hyperprolactinemia, which is known as the "stalk effect". However, the relationships between hyperprolactinemia and the radiographic characteristics of the tumor that affects the pituitary stalk have not been well characterized. We aimed to identify the differences in the clinical and radiographic characteristics of patients with NFPA, with and without hyperprolactinemia. METHODS: We enrolled 107 patients with NFPA and allocated them to hyperprolactinemia and non-hyperprolactinemia groups using two different cut-off values: (1) the upper limit of the normal reference range, adjusted for sex and menopausal status, and (2) the upper quartile across the cohort, and compared their clinical and radiographic characteristics. These analyses were conducted to clarify the relationship between the "stalk effect" and the postoperative change in antidiuretic hormone secretion. RESULTS: The specific radiographic characteristics of the patients included the presence of a cystic or hemorrhagic tumor and the presence of pituitary stalk deviation, which were more frequent in the patients with hyperprolactinemia. Interestingly, the incidence of postoperative transient diabetes insipidus was statistically significantly higher in the hyperprolactinemia group (≥40 ng/mL) and in the group with radiologic evidence of stalk deviation, which were shown to be independent risk factors on multivariate analysis. CONCLUSION: The presence of a "stalk effect" was associated with a higher risk of postoperative transient diabetes insipidus, reflecting perioperative pituitary stalk dysfunction following NFPA surgery, especially in patients with serum prolactin concentrations ≥40 ng/mL and radiologic evidence of stalk deviation.

Bold-S Signs on Computed Tomography Angiography Are Sensitive Markers for Diagnosing Subcortical Hemorrhage Due to Dural Arteriovenous Fistulae on Emergent Admission.

An optimal treatment strategy for subcortical hematomas caused by dural arteriovenous fistulae (dAVF) is important because of the high rebleeding rate. However, it is very difficult to diagnose that on admission. Therefore, an early sensitive predictive marker for subcortical hemorrhage caused by dAVF is necessary, especially during the first contact on admission. S-shaped dilated vessels around the hematoma (bold-S sign) on computed tomography angiography (CTA) performed during admission could be one such marker. Herein, we evaluated the characteristics of these vessels. Among 273 patients with intracerebral hemorrhage between April 2012 and March 2020, 67 patients with subcortical hematomas who underwent CTA on admission without arteriovenous malformations were included. The patients in the dAVF group (n = 7) showed fewer disturbances in consciousness, milder neurological deficits, and more frequent seizures than patients without dAVF (without dAVF group, n = 60). All patients in the dAVF group had dilated S-shaped vessels (2.59 ± 0.27 mm) around the hematomas, and only 20% of the patients in the without dAVF group had these vessels (1.69 ± 0.22 mm). The ratio of the ipsilateral S-shaped/contralateral largest vessels was 1.80 ± 0.29 in the dAVF group and 1.07 ± 0.16 in the group without dAVF. We called the dilated S-shaped vessels the "bold-S sign," with a cutoff ratio of 1.5. Bold-S sign findings are novel and help in diagnosing subcortical hematomas caused by dAVF on admission.

Effects of Microvascular Decompression on Pain Relief and Quality of Life in Late Elderly Patients with Trigeminal Neuralgia.

Pharmacotherapy is frequently selected over surgical interventions for late elderly patients with trigeminal neuralgia (TN). However, medication may affect these patients' activities of daily living (ADL). Hence, we investigated the effect of the surgical treatment of TN on ADL in older patients. This study included 11 late elderly patients >75 years old and 26 nonlate elderly patients who underwent microvascular decompression (MVD) for TN at our hospital from June 2017 to August 2021. We evaluated pre- and postsurgical ADL using the Barthel Index (BI) score, side effects of antineuralgic drugs, the BNI pain intensity score, and perioperative medication. The BI score of late elderly patients significantly improved postoperatively, particularly in transfer (pre: 10.5; post: 13.2), mobility (pre: 10; post: 12.7), and feeding (pre: 5.9 points; post: 10 points). Additionally, antineuralgic drugs caused preoperative disturbances of transfer and mobility. Trends of a longer disease duration and frequent occurrence of side effects were observed in all patients in the elderly group, compared to only 9 out of 26 patients in the younger group (100% vs. 35%, p = 0.0002). In addition, drowsiness was observed more frequently in the late elderly group (73% vs. 23%, p = 0.0084). However, the change in scores indicating improvement after surgery was significantly greater in the late elderly group, although both pre- and postoperative scores were higher in the nonlate elderly group (11.4 ± 1.9 vs. 6.9 ± 0.7, p = 0.027). Surgical treatment can improve older patients' ADL because it relieves pain and facilitates discontinuation of antineuralgic drugs. Consequently, MVD can be positively recommended for older patients with TN if general anesthesia is acceptable.

Safety and optimal management of hepatic arterial infusion chemotherapy after pancreatectomy for pancreatobiliary cancer.

OBJECTIVE: The purpose of this study was to assess the safety of hepatic arterial infusion (HAI) chemotherapy after pancreatectomy for pancreatobiliary cancer. MATERIALS AND METHODS: The study subjects comprised 51 patients treated with HAI chemotherapy after pancreatectomy: 31 patients had undergone pancreaticoduodenectomy or total pancreatectomy and 20 patients had undergone distal pancreatectomy. HAI chemotherapy was performed for the prevention of liver metastasis in 42 patients and for the treatment of metastases in nine patients. HAI chemotherapy consisted of 5-fluorouracil administered at 1000 mg/m(2)/wk. After each cycle of chemotherapy, the patency of the hepatic artery and complications were evaluated using digital subtraction angiography and CT arteriography via the catheter-port system. RESULTS: Placement of the catheter-port system was successful in all patients. Hepatic arterial occlusion and severe asymptomatic hepatic artery stenosis were observed in one patient (2%) and 10 patients (19.6%), respectively. The hepatic arterial patency rate tended to be lower in the group who underwent pancreaticoduodenectomy or total pancreatectomy (71%) than in the group who underwent distal pancreatectomy (90%), although the difference was not significant (p = 0.107). Four asymptomatic patients with hepatic artery stenosis restarted HAI chemotherapy after a 4-week withdrawal of treatment. In three patients (5.9%), all of whom had undergone pancreaticoduodenectomy, liver abscess or biloma developed synchronously with asymptomatic hepatic artery stenosis. All three cases were cured by percutaneous abscess drainage or conservative management. The tumor response rate in the treatment of liver metastases was 44.4%. The hepatic recurrence rate with adjuvant therapy was 7.1%. CONCLUSION: Our findings suggest that HAI chemotherapy after pancreatectomy is feasible and tolerable, although caution is required particularly after pancreaticoduodenectomy or after total pancreatectomy.

CT Findings and Treatment Outcomes of Ground-Glass Opacity Predominant Lung Cancer After Stereotactic Body Radiotherapy.

BACKGROUND: Stereotactic body radiotherapy (SBRT) has been rapidly evolving and increasingly performed in patients with ground-glass opacity (GGO) predominant lung cancer (GGOp-LC). PURPOSE: To evaluate early-phase CT findings of GGOp-LC after SBRT. MATERIALS AND METHODS: Patients with GGOp-LC staged as cTis-2bN0M0 treated with SBRT were retrospectively identified. The CT images were analyzed using radiologists' interpretation and CT-density histograms. Long-term treatment outcomes were also assessed. RESULTS: This study evaluated 126 patients with 133 cases of GGOp-LC, comprising GGOp-LC with pure GGO (pureGGO-LC) (n = 31) and part-solid tumors (partsolid-LC) (n = 102). The median follow-up duration was 64.3 months (range, 10.8-178.9 months). Most GGOp-LC cases were interpreted as stable disease at 1 and 3 months after SBRT (96% [125/130] and 85% [62/73], respectively). However, the solid component was often interpreted as progressive disease (42% [34/82] and 60% [29/48], respectively). The GGO component was interpreted as denser in 47% (61/130) and 86% (63/73) of cases, respectively. For 25 evaluable pureGGO-LC cases at 3 months, the median tumor density values increased over time (P < .001). For 48 evaluable partsolid-LC cases at 3 months, the median areas of CT-density ≥ -160 HU increased over time (P < .001). The 5-year overall survival for GGOp-LC patients was 78.0%. No local or regional recurrence were observed. CONCLUSION: Clinical outcomes of SBRT for GGOp-LC were excellent, without local or regional recurrence. In the interpretation of early-phase follow-up CT scans of GGOp-LC after SBRT, it should be noted that most GGOp-LC remains stable disease, solid component increases in size, and GGO component is denser.

A novel 8.57-kb deletion of the upstream region of PRKAR1A in a family with Carney complex.

Carney complex (CNC) is a rare hereditary syndrome that involves endocrine dysfunction and the development of various types of tumors. Chromosome 2p16 and PRKAR1A on chromosome 17 are known susceptibility loci for CNC. Here we report a mother and son with CNC caused by an 8.57-kb deletion involving the transcription start site and non-coding exon 1 of PRKAR1A. The proband is a 28-year-old male with bilateral large-cell calcified Sertoli cell testicular tumors and pituitary adenoma. Comprehensive genomic profiling for cancer mutations using Foundation One CDx failed to detect any mutations in PRKAR1A in DNA from the testicular tumor. Single-nucleotide polymorphism array analysis of the proband's genomic DNA revealed a large deletion in the 5' region of PRKAR1A. Genomic walking further delineated the region an 8.57-kb deletion. A 1.68-kb DNA fragment encompassed by the deleted region showed strong promoter activity in a NanoLuc luciferase reporter assay. The patient's mother, who is suffering from recurrent cardiac myxoma, a critical sign for CNC, carried an identical deletion. The 8.57-kb deleted region is a novel lesion for CNC and will facilitate molecular diagnosis of the disease.

Analysis of onset and clinical characteristics in Japanese patients with infantile hemangioma.

Infantile hemangioma (IH) is a common benign tumor during infancy, although the detailed mechanism behind it has not been fully elucidated. Based on previous studies, we hypothesized that formation of hemangioma might be triggered by secondary physiological events (perinatal hypoxia or mechanical stress during delivery) in patients carrying germline risk mutations. We aimed to clarify the mechanism by evaluating whether head and neck lesions were more frequent in patients in who IH appeared after birth compared with those in who it was present at birth. Clinical data of 62 lesions in 51 patients with IH were collected. All patients were analyzed for correlation of onset with gender, localization, family histories, gestational age, birth weight, and clinical subtypes. Distribution of lesions on the head and neck was slightly more frequent in the after-birth IH group, compared with those with IH present at birth, but without significant difference (47.6% vs. 40.0%, p = 0.32). On the other hand, the ratio of superficial and deep type IH at birth was significantly altered compared with that in IH after birth (19:0 vs. 26:7, p = 0.039). In addition, IHs appearing after birth tended to more commonly have multiple lesions than those with IH present at birth, with statistically significant difference (25.8% vs. 0%, p = 0.0164). There may therefore be different triggers for IHs at birth and IH after birth. Further studies with greater number of patients are necessary to validate these findings.

Additional Embolization for Intra-aneurysmal Blood Flow Resumption after Stent-assisted Embolization of Cerebral Aneurysms.

Objective: Stent-assisted aneurysmal embolization (SAAE) is an effective treatment for aneurysms with a low risk of recurrence. In rare cases, retreatment is necessary due to recanalization of blood flow into the aneurysm. However, only a few studies have reported on retreatment. We examined the efficacy and complications of stent-assisted aneurysm embolization for large or wide-neck aneurysms at our hospital. Methods: Between July 2010 and June 2018, 293 patients underwent stent-assisted aneurysm embolization at our hospital. Among them, 12 (2 women, 10 men, mean age: 62 years) needed retreatment. We evaluated the initial treatment of these 12 patients, and the methods and results of their retreatment. Results: Six of the 12 retreated patients were treated using the simple technique. It was possible to treat nine patients (75%) without placing new stents, but three needed additional stents. We were able to guide the microcatheter into the aneurysm using the trans-cell technique even with two overlapping stents. We achieved complete embolism in seven patients (58%), and remnants were observed in the neck in five (42%) patients. No complications were associated with our surgery. We were able to perform follow-up for 10 patients and there was no recurrence. Conclusion: Embolization should be considered in recurrent cases after the initial stent-assisted coil embolization. We achieved good results and reduced the recurrence rate by selecting the appropriate treatment in each case.

Chronological dynamic changes in cortico-subcortical imbalance of cerebral blood flow in a boy with CAPOS syndrome.

BACKGROUND: Cerebellar ataxia, Areflexia, Pes cavus, Optic atrophy and Sensorineural hearing loss (CAPOS) syndrome is a known ATP1A3-related disorder, but little has been elucidated regarding its pathophysiology. We now report two new patients, a Japanese boy and his mother with a pathogenic mutation (c.2452G>A) in ATP1A3, who were diagnosed with CAPOS syndrome. METHODS: After febrile illnesses at 7 months of age, and again at 22 months of age, the boy had a reduced level of consciousness, truncal ataxia and eye movement-disorders. The patient's 32-year-old mother may have experienced an episode of acute encephalopathy in her childhood and sustained sensorineural hearing loss. In the present study, we demonstrated chronological dynamic changes in cerebral blood flow (CBF) in the son, using serial single-photon emission computed tomography (SPECT). RESULTS: The serial CBF-SPECT findings using statistical methods showed progressive hyperperfusion in the frontal lobes, basal ganglia and thalamus, and hypoperfusion in the occipital and temporal lobes during the acute and subacute phases. Thereafter, the dynamic changes of CBF improved in the chronic but hypoperfusion in thalamus appeared to the chronic phase. CONCLUSION: The abnormal cortico-subcortical CBF may contribute to an acute encephalopathy-like condition in the acute stage of CAPOS syndrome. CAPOS syndrome is not often reported, and is possibly an under-recognized syndrome in clinically mild cases.

Influence of conditioning regimens and stem cell sources on donor-type chimerism early after stem cell transplantation.

We retrospectively analyzed very early chimerism before and ongoing neutrophil engraftment (days 7, 14, 21, 28) and investigated the influence of conditioning regimens and stem cell sources on donor-type chimerism in 59 Japanese patients who had received allogeneic hematopoietic stem cell transplantation. The percentage of donor-type chimerism increased before engraftment in all patients who achieved engraftment. The average percentage of donor-type chimerism in patients who had received reduced-intensity stem cell transplantation (RIST) with total body irradiation (TBI) was significantly higher than that in patients who had received RIST without TBI (98.8% vs 87.5% on day 21, P<0.01; 99.3% vs 84.3% on day 28, P<0.01). The average percentage of donor-type chimerism after peripheral blood stem cell transplantation was significantly higher than that after bone marrow transplantation on day 7 (81.5% vs 43.1%, P<0.01), and the average percentage of donor-type chimerism after cord blood transplantation was significantly lower on day 14 (55.8% vs 84.8%, P<0.05). Compared with the average percentage of donor-type chimerism in patients who achieved engraftment with each stem cell source, a notable decrease in donor-type chimerism was observed in patients who failed to achieve engraftment. This study suggests that differences in conditioning regimens and stem cell sources should be taken into account when considering donor-type chimerism.

A case of uterine torsion concurrent with a ruptured ovarian endometrial cyst.

The patient was a 54-year-old female presented with severe abdominal swelling and intermittent pain. On emergent CT, massive ascites with thickened peritoneum and intra-pelvic cystic mass approximately 20 cm in diameter were observed. The cyst wall showed redundant irregular shape. The uterus was enlarged with intramural mass located at its right anterior fundus. MRI showed this pelvic cyst as high intensity on T1-weighted images, so that it was suspected as an endometrial cyst. The operation revealed the rupture of endometrial cyst and the uterine torsion of 180° around the long axis. Retrospectively, the X-shaped configuration of the upper vagina was observed on MRI and both the adnexa, including right ovarian cyst, were connected to the opposite side ovarian vein. Torsion of a non-gravid uterus is rare. In this case, the uterine torsion seemed to be caused by enlarged uterine body and ruptured endometrial cyst. Radiologists should be aware of this potential complication of huge ovarian mass and enlarged uterus and its appearance.

Adjuvant Hepatic Arterial Infusion Chemotherapy After Resection for Pancreatic Cancer Using Coaxial Catheter-Port System Compared with Conventional System.

PURPOSE: Previous reports have shown the effectiveness of adjuvant hepatic arterial infusion chemotherapy (HAIC) in pancreatic cancer. However, percutaneous catheter placement is technically difficult after pancreatic surgery. The purpose of this study was to evaluate the feasibility and outcome of HAIC using a coaxial technique compared with conventional technique for postoperative pancreatic cancer. MATERIALS AND METHODS: 93 consecutive patients who received percutaneous catheter-port system placement after pancreatectomy were enrolled. In 58 patients from March 2006 to August 2010 (Group A), a conventional technique with a 5-Fr indwelling catheter was used and in 35 patients from September 2010 to September 2012 (Group B), a coaxial technique with a 2.7-Fr coaxial catheter was used. RESULTS: The overall technical success rates were 97.1 % in Group B and 86.2 % in Group A. In cases with arterial tortuousness and stenosis, the success rate was significantly higher in Group B (91.7 vs. 53.8 %; P = 0.046). Fluoroscopic and total procedure times were significantly shorter in Group B: 14.7 versus 26.7 min (P = 0.001) and 64.8 versus 80.7 min (P = 0.0051), respectively. No differences were seen in the complication rate. The 1 year liver metastasis rates were 9.9 % using the conventional system and 9.1 % using the coaxial system (P = 0.678). The overall median survival time was 44 months. There was no difference in the survival period between two systems (P = 0.312). CONCLUSIONS: The coaxial technique is useful for catheter placement after pancreatectomy, achieving a high success rate and reducing fluoroscopic and procedure times, while maintaining the safety and efficacy for adjuvant HAIC in pancreatic cancer.

Time-lapse Raman imaging of osteoblast differentiation.

Osteoblastic mineralization occurs during the early stages of bone formation. During this mineralization, hydroxyapatite (HA), a major component of bone, is synthesized, generating hard tissue. Many of the mechanisms driving biomineralization remain unclear because the traditional biochemical assays used to investigate them are destructive techniques incompatible with viable cells. To determine the temporal changes in mineralization-related biomolecules at mineralization spots, we performed time-lapse Raman imaging of mouse osteoblasts at a subcellular resolution throughout the mineralization process. Raman imaging enabled us to analyze the dynamics of the related biomolecules at mineralization spots throughout the entire process of mineralization. Here, we stimulated KUSA-A1 cells to differentiate into osteoblasts and conducted time-lapse Raman imaging on them every 4 hours for 24 hours, beginning 5 days after the stimulation. The HA and cytochrome c Raman bands were used as markers for osteoblastic mineralization and apoptosis. From the Raman images successfully acquired throughout the mineralization process, we found that ß-carotene acts as a biomarker that indicates the initiation of osteoblastic mineralization. A fluctuation of cytochrome c concentration, which indicates cell apoptosis, was also observed during mineralization. We expect time-lapse Raman imaging to help us to further elucidate osteoblastic mineralization mechanisms that have previously been unobservable.