Evaluation of the Effects of Cultured Bone Marrow Mesenchymal Stem Cell Infusion on Hepatocarcinogenesis in Hepatocarcinogenic Mice With Liver Cirrhosis.

OBJECTIVES: Liver transplantation remains the only curative therapy for decompensated liver cirrhosis. However, it has several limitations, and not all patients can receive liver transplants. Therefore, liver regenerative therapy without liver transplantation is considered necessary. In this study, we attempted minimally invasive liver regenerative therapy by peripheral vein infusion of bone marrow-derived mesenchymal stem cells (BMSCs) cultured from a small amount of autologous bone marrow fluid and evaluated the effects of BMSCs on hepatocarcinogenesis in a mouse model. METHODS: C57BL/6 male mice were injected intraperitoneally with N-nitrosodiethylamine once at 2 weeks of age, followed by carbon tetrachloride twice a week from 6 weeks of age onwards, to create a mouse model of highly oncogenic liver cirrhosis. From 10 weeks of age, mouse isogenic green fluorescent protein-positive BMSCs (1.0 × 106/body weight) were infused once every 2 weeks, for a total of 5 times, and the effects of frequent BMSC infusion on hepatocarcinogenesis were evaluated. RESULTS: In the histologic evaluation, no significant differences were observed between the controls and BMSC-administered mice in terms of incidence rate, number, or average size of foci and tumors. However, significant suppression of fibrosis and liver injury was confirmed in the group that received BMSC infusions. DISCUSSION: Considering that BMSC infusion did not promote carcinogenesis, even in the state of highly oncogenic liver cirrhosis, autologous BMSC infusion might be a safe and effective therapy for human decompensated liver cirrhosis.

De novo sequencing of highly modified therapeutic oligonucleotides by hydrophobic tag sequencing coupled with LC-MS.

Correct sequences are prerequisite for quality control of therapeutic oligonucleotides. However, there is no definitive method available for determining sequences of highly modified therapeutic RNAs, and thereby, most of the oligonucleotides have been used clinically without direct sequence determination. In this study, we developed a novel sequencing method called 'hydrophobic tag sequencing'. Highly modified oligonucleotides are sequenced by partially digesting oligonucleotides conjugated with a 5'-hydrophobic tag, followed by liquid chromatography-mass spectrometry analysis. 5'-Hydrophobic tag-printed fragments (5'-tag degradates) can be separated in order of their molecular masses from tag-free oligonucleotides by reversed-phase liquid chromatography. As models for the sequencing, the anti-VEGF aptamer (Macugen) and the highly modified 38-mer RNA sequences were analyzed under blind conditions. Most nucleotides were identified from the molecular weight of hydrophobic 5'-tag degradates calculated from monoisotopic mass in simple full mass data. When monoisotopic mass could not be assigned, the nucleotide was estimated using the molecular weight of the most abundant mass. The sequences of Macugen and 38-mer RNA perfectly matched the theoretical sequences. The hydrophobic tag sequencing worked well to obtain simple full mass data, resulting in accurate and clear sequencing. The present study provides for the first time a de novo sequencing technology for highly modified RNAs and contributes to quality control of therapeutic oligonucleotides. Copyright © 2016 John Wiley & Sons, Ltd.

sLORETA-qm for interictal MEG epileptic spike analysis: comparison of location and quantity with equivalent dipole estimation.

OBJECTIVE: To determine whether quantitative modification of a standardised low-resolution brain electromagnetic tomography (sLORETA-qm) could be used as a reliable tool for quantitative analysis of magnetoencephalography (MEG) for analysis of the interictal epileptic spike. To verify the performance of sLORETA-qm, magnetic source location and quantity were compared with the equivalent current dipole (ECD) method. METHODS: A total of 50 sources from 10 patients with epilepsy were obtained. Analyses were performed after the MEG data were 3-70 Hz band-pass filtered. Time points for analysis were selected referring to waveform patterns and the isofield contour map. With the same spherical model, source estimation was conducted with two methods of analysis: ECD and sLORETA-qm. Distance from the centre of the spherical model and intensities were compared between the methods. RESULTS: There were no significant differences between the methods in the distance from the spherical model (paired t-test, p=0.8761). Source intensities between the methods were strongly correlated (Spearman's Rho=0.9803, p<0.001). CONCLUSIONS: sLORETA-qm was closely correlated with ECD concerning point source location and quantity in analysis of the interictal epileptic spike. SIGNIFICANCE: sLORETA-qm is a reliable quantifiable method without arbitrariness for analysis of the interictal epileptic spike.

Significant relationship between local angle at fused segments and C2-7 angle: Average duration of longer than 20 years after anterior cervical discectomy and fusion.

BACKGROUND: The authors have focused their attention to the radiological durability of cervical sagittal alignment after anterior cervical discectomy and fusion (ACDF) using autologous bone grafting. MATERIALS AND METHODS: Among the patients who underwent ACDF with trans-unco-discal (TUD) approach between 1976 and 1997, 22 patients (16 males and 6 females) made return visits for a clinical evaluation. Patients with trauma or previously treated by anterior cervical fusion or by posterior decompression were excluded from the present study. Clinical evaluation included adjacent segment degeneration (ASD), osseous fusion, local angle at the fused segments and C2-7 angle of cervical spine. RESULTS: The duration after ACDF ranged from 13 to 34 years with an average of 21.3 ± 7.0 years. A single level fusion was done on 8 patients, 2 levels on 11 patients, 3 levels on 2 patients, and 4 levels on 1 patient. Imaging studies indicated that 12 of the 22 patients (54.5%) were graded as having symptomatic ASD. Osseous bony fusion at ACDF was recognized in all cases. None of the patients demonstrated kyphotic malalignment of the cervical spine. Average degrees of local angle at the fused segments and the C2-7 angle were 7.06 and 17.6, respectively. Statistical analysis indicated a significant relationship between the local at the fused segments and C2-7 angles. CONCLUSIONS: Sagittal alignment of the cervical spine was durable long after ACDF when the local angle at the fused segments was well stabilized.

Normal variation of diffusion tensor parameters of the spinal cord in healthy subjects at 3.0-Tesla.

AIMS: The purposes of the present study were to clarify the normal variation and to determine the normal reference values of diffusion tensor (DT) parameters (mean diffusivity [MD] and fractional anisotropy [FA]) of the spinal cord in single-shot fast spin-echo-based sequence at 3.0-Tesla (3T). MATERIALS AND METHODS: Thirty healthy subjects (mean age = 44.2 years, range = 20-72 years) were enrolled for this study. Mean values of MD and FA in six spinal levels (C2/3, C3/4, C4/5, C5/6, C6/7, and C7/Th1) were measured. Mean values, variances, and distributions of the MD and FA in each spinal level were analyzed. Age-dependent change of MD and FA as well as correlation between MD and FA was also analyzed. RESULTS: At all spinal levels, the values can be considered to be Gaussian distribution in MD but not in FA. A significant statistical negative correlation was observed between aging and the values of MD (r = 0.429, P = 0.018), but insignificant between the values of FA (P = 0.234). A slight significant statistical negative correlation was observed between the values of MD and FA (r = 0.156, P = 0.037). One way repeated measures analysis of variance indicated the significant difference between the spinal levels in both MD (P = 0.003) and FA (P < 0.0001). CONCLUSIONS: The analyzed data in the present study would be helpful for comparison when investigating the spinal condition of spinal disorders.

Psychotic-like experiences are associated with suicidal feelings and deliberate self-harm behaviors in adolescents aged 12-15 years.

OBJECTIVE: Psychotic disorders are a significant risk factor for suicide, especially among young people. Psychotic-like experiences (PLEs) in the general population may share an etiological background with psychotic disorders. Therefore, the present study examined the association between PLEs and risk of suicide in a community sample of adolescents. METHOD: Psychotic-like experiences, suicidal feelings, and self-harm behaviors were studied using a self-report questionnaire administered to 5073 Japanese adolescents. Depression and anxiety were evaluated using the 12-item General Health Questionnaire (GHQ). RESULTS: The presence of PLEs was significantly associated with suicidal feelings (OR = 3.1, 95% CI = 2.2-4.5) and deliberate self-harm behaviors (OR = 3.1, 95% CI = 2.0-4.8) after controlling for the effects of age, gender, GHQ-12 score, victimization, and substance use. Suicidal feelings and behaviors were more prevalent in subjects with a greater number of PLEs. CONCLUSION: Psychotic-like experiences may increase the risk of suicidal problems among adolescents.

Endoscopic ultrasound-guided fine needle aspiration biopsy for splenic tumor: a case series.

Splenic tumors are occasionally found in clinical practice but the diagnosis is often difficult if only serologic and imaging tests are used. Therefore, pathologic sampling is required in such cases. Endoscopic ultrasonography (EUS) provides a good image of the spleen through the gastric wall, and a transgastric EUS-guided fine needle aspiration (EUS-FNA) biopsy may be easier than the percutaneous approach. Furthermore, a large-gauge needle may raise the capability of EUS-FNA for the histopathologic diagnosis. The aim of this study was to evaluate the yield of EUS-FNA using a large-gauge needle for a splenic tumor. Five patients with splenic tumor were subjected to EUS-FNA with a 19-gauge needle to obtain histopathologic materials. A pathologic sample was obtained in all cases, and the diagnoses were lymphoma (n = 2), sarcoidosis (n = 2), and inflammatory pseudotumor (n = 1). EUS-FNA using a 19-gauge needle is safe and useful for the diagnosis of splenic tumors.

Quantitative analysis of MEG using modified sLORETA for clinical application.

OBJECTIVE: To determine whether standardised low-resolution brain electromagnetic tomography modified for a quantifiable method (sLORETA-qm) can be used for quantitative analysis in magnetoencephalography (MEG). METHODS: Somatosensory evoked fields (SEFs) were obtained from 10 hemispheres of five healthy volunteers stimulated on the median nerve at 0.75, 1.0, 1.25, 1.5, 1.75 and 2.0 x threshold of thenar muscle twitch (TMT). N20 m intensity changes were analysed quantitatively using sLORETA-qm. Then, SEFs were measured with stimulation on the median nerve at 1.5 x TMT from 47 hemispheres in 24 subjects. sLORETA-qm intensity and the equivalent current dipole (ECD) moment of N20 m were calculated, and relationships between the values were evaluated. RESULTS: sLORETA-qm intensity increased linearly with stimulus intensity between 0.75 and 1.5 x TMT, and tended to reach a plateau or decrease at higher stimulus intensities. The distribution of sLORETA-qm intensity after natural logarithmic transformation was normal and a close correlation was found between the ECD moment and sLORETA-qm intensity (r(s)=0.91, p<0.001). CONCLUSIONS: The results of this study focusing on N20 m suggested that sLORETA-qm is reliable for quantitative analysis of MEG as well as ECD models. SIGNIFICANCE: sLORETA-qm appears promising for quantitative analyses of MEG for which ECD models are inappropriate.

Atypical spinal dural arteriovenous fistula with supply from the lateral sacral artery.

We report a dural arteriovenous fistula (AVF) that developed at a site on the midline dorsal surface of the dura mater that had been damaged by repeated lumbar punctures. A 61-year-old male patient had undergone repeated lumbar punctures and discectomy for severe lumbago 40 years before the present admission. After surgery, the lumbago symptoms resolved. However, 30 years after the operation, he started to experience dysaesthesia, motor weakness in both legs, and urinary disturbance. Physical examination revealed bilateral leg weakness, diminished deep tendon reflexes in the patellar and Achilles tendons bilaterally, and decreased superficial sensation below L1. Magnetic resonance imaging revealed swelling with intramedullary high intensity and multiple flow voids around the conus and spinal cord on T(2)-weighted images, and adhesive arachnoiditis. Spinal angiography revealed an AVF between the left lateral sacral artery and the S1 radicular vein at the site of the previous operation. Surgery was conducted to carry out excision of the dural AVF at the shunting point, the arterialized intradural vein, and lysis of the arachnoiditis. This case of dural AVF may have been caused by repeated lumbar punctures.

Endoscopic ultrasound-guided fine-needle aspiration biopsy for lymphadenopathy of unknown origin.

BACKGROUND AND STUDY AIMS: The diagnosis of mediastinal and intra-abdominal lymphadenopathy is sometimes difficult, especially in patients who have no other primary lesions. Lymphoma is one of the main causes of this condition. Endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) is a safe and accurate diagnostic procedure for lesions surrounding the gastrointestinal tract. However, diagnosing lymphoma using the EUS-FNA technique remains a diagnostic challenge, due to limitations in the amount of material sampled. The aim of the present study was to evaluate the yield of EUS-FNA biopsy (EUS-FNAB) using a large-gauge needle in patients with mediastinal and intra-abdominal lymphadenopathy of unknown origin, especially in relation to subclassification of the lymphomas. PATIENTS AND METHODS: Consecutive patients with mediastinal and intra-abdominal lymphadenopathy of unknown origin who were referred between October 2003 and March 2005 were enrolled in the study. EUS-FNAB was carried out using a 19-gauge needle, passing through the esophageal, gastric, and duodenal walls. Pathological diagnoses were made on the basis of histological findings, including immunopathological staining. RESULTS: A total of 104 patients were included in the study. The locations of the lymph nodes were mediastinal in 50 patients, intra-abdominal in 48 patients, and both mediastinal and intra-abdominal in six patients. The diagnoses made using EUS-FNAB were lymphoma (n = 48), metastasis (n = 16), and benign/reactive (n = 40). The overall accuracy of EUS-FNAB for unknown lymphadenopathy was 98 %, and it was possible to classify the lymphomas in accordance with the World Health Organization classifications in 88 % of cases. No serious complications occurred with the procedure. CONCLUSIONS: Open thoracic surgery, laparotomy, and other invasive diagnostic procedures such as mediastinoscopy and laparoscopy can now be avoided, as EUS-FNAB is potentially a safe and accurate tool for diagnosing unknown lymphadenopathy, including lymphoma.

Evidence of two dimensionality in quasi-one-dimensional cobalt oxides.

The quasi-one-dimensional (Q1D) cobalt oxides A(N + 2)Co(n + 1)O(3n + 3) (A = Ca, Sr, and Ba, n = 1 - infinity) were investigated by muon-spin spectroscopy under applied pressures of up to 1.1 GPa. The relationship between the onset Néel temperature T(on)(N) and the interchain distance (d(ic)), which increases monotonically with n, is well fitted by the formula T(N)/T(N,0) = (1 - d(ic)/d(ic,o)(beta), here for T(on)(N) approximately 100 K for Ca(3)Co(2)O(6) (n = 1) and approximately 15 for BaCoCoO(3) (n = infinity at ambient P. The T(on)(N) - d(ic) curve also predicts a large dependence of Y(N) for the compounds with n > or = 5, i.e., in the vicinity of , while the compounds show only a very small effect. Indeed, our high-pressure mu(+) results show that of BaCoO(3) is enhanced by with a slope of 2.2 K(Gpa), whereas no detectable changes by P for both Ca(3)Co(2)O(6) and Sr(4)Co(3)O(9) (n = 2). This clearly confirms the role of the 2D-antiferromagnetic interaction on T(on)(N) in the Q1D cobalt oxides.

Hemangioblastoma of hippocampus without von Hippel-Lindau disease: case report and review of literature.

A rare case of hemangioblastoma located in the region of hippocampus is reported. A 27-year-old female presented with a single episode of generalized convulsion. The vascular and cherry red color hemangioblastoma was resected by a temporo-zygomatic approach. There has been no recurrence of tumor at a follow-up of 11 years.

Correlations of hemoglobin index (IHb) of gastric mucosa with Helicobacter pylori (H. pylori) infection and inflammation of gastric mucosa.

BACKGROUND: Helicobacter pylori (H. pylori) infection causes various gastric diseases, among them H. pylori-associated gastritis characterized by diffuse redness of the gastric mucosa. The haemoglobin index (IHb) of the fundic mucosa is an objective parameter of the extent of mucosal redness, but it is unclear whether or not IHb can be used as a diagnostic marker for H. pylori infection. The purpose of this investigation was to evaluate the correlations between IHb of the fundic mucosa and H. pylori infection, inflammatory cell infiltration, and inflammatory mediator production. METHODS: IHb of the fundic mucosa was measured in 108 patients with various gastric diseases (group 1), and values were compared between H. pylori-positive and H. pylori-negative patients. Fifteen patients with H. pylori infection from group 1 underwent H. pylori eradication therapy and IHb was measured before and after treatment. Both IHb and inflammatory cell infiltration were assessed in 61 patients (group 2). In 31 patients from group 2, the expression of interleukin (IL)-8 and inducible nitric oxide synthase (iNOS) messenger RNA (mRNA) was assayed in gastric biopsy specimens by the reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: IHb levels were significantly higher in H. pylori-positive patients than in H. pylori-negative patients (P < 0.001). IHb was decreased at one month after the eradication of H. pylori (P < 0.001). IHb was higher in patients with infiltration by both mononuclear cells and neutrophils (P < 0.001). There was a significant correlation between the IHb level and the expression of IL-8 mRNA (P < 0.001), as well as between IHb and iNOS mRNA expression (P < 0.05). CONCLUSIONS: There were significant correlations between IHb of the gastric mucosa and H. pylori infection, inflammatory cell infiltration, and IL-8/iNOS mRNA expression, suggesting that IHb is a reliable marker of H. pylori infection for use during follow-up endoscopy after H. pylori eradication therapy.

Three-dimensional rotation venography using the digital subtraction angiography unit with a flat-panel detector: usefulness for the transtemporal/transtentorial approaches.

We obtained the venograms using the two-dimensional digital subtraction angiography (2D DSA) images and three dimensional rotation venography (3D RV) images and investigated the potential usefulness of the 3D RV compared with venograms of 2D DSA using the newly developed three-dimensional rotation angiography unit with a flat-panel detector (FPD). This study included 26 sides (11 left, 15 right) in 20 cases (4 males and 16 females) who underwent radiographic examination for management of intracranial tumors and vascular diseases between May 2003 and December 2003. Each patient underwent diagnostic angiography performed on a DSA unit with a FPD. In all patients, the 2D DSA images, including anteroposterior view and lateral view of the carotid artery, were obtained in two stereoscopic views. The 3D RV was used to produce volume-rendered images. Two neuroradiologists investigated the venous configuration of 3D RV compared with that of 2D DSA about the relationship of the venous drainage system on the temporal lobe according to Guppy et al.'s classification. Twenty-four sides of the 26 sides enabled the precious visualization on 3D RV images. In investigation of 2D DSA, 9 sides (37.5%) were classified into type A, 13 (54.2%) into type B, two (8.3%) into type C, and no sides into types D, E, and F. In investigation of 3D RV images, 10 sides (41.7%) were classified into type A, 9 (37.5%) into type B, 1 (4.2%) into type C, 2 (8.3%) into type E, and 2 (8.3%) into type F. Seven of 24 sides demonstrated discrepancy in results between 2D DSA and 3D RV. The 3D RV could be performed by setting the adequate delay in between the injection of the contrast material and starting time of third rotation to acquire the opacified images. In Guppy et al.'s classification, the 3D RV images could demonstrate the precious venous drainage including the venous lakes with use of multiple views and variable reconstruction compared with 2D DSA. Our DSA system with FPD could provide good 3D RV images. These images are very useful for the skull-base surgery because we can understand the three-dimensional vascular anatomy preoperatively.

Lift-up laminoplasty for myelopathy caused by ossification of the posterior longitudinal ligament of the cervical spine.

BACKGROUND AND AIMS: We have utilized lift-up laminoplasty to treat patients with myelopathy caused by ossification of the posterior longitudinal ligament (OPLL) of the cervical spine. The preliminary surgical outcome with computer-assisted morphological assessment is presented. MATERIAL AND METHODS: The surgical technique of lift-up laminoplasty includes standard posterior exposure of the cervical spine, en-bloc laminectomy, and expansion of the cervical canal by lift-up of the laminae with custom-designed hydroxyapatite laminar spacers and stabilization of the laminae using titanium miniplates. From 1998 to 2003, 10 consecutive patients with cervical myelopathy secondary to OPLL have been treated with this method and comprehensively evaluated. Care was taken to tailor the treatment to individual patients by using different sizes of spacers to adjust the degree of expansion depending on the amount of stenosis of the cervical spine. The degree of expansion of the cervical canal was altered by design, based on the preoperative imaging simulation. RESULTS: Preliminary surgical outcome, evaluated at 6 months after surgery, revealed a significant improvement of neurological function. Image analysis revealed that the cervical canals were significantly expanded, with a mean reduction of 13.1% in the stenosis ratio. Lift-up laminoplasty was effective in the treatment of patients with myelopathy secondary to cervical OPLL, and the amount of expansion could be individually adjusted at the discretion of the surgeon. CONCLUSION: Although analysis with a larger population and a longer follow-up period needs to be undertaken, our method of lift-up laminoplasty appears to be a viable choice among standard posterior cervical approaches for cervical OPLL.

Extra-axial subarachnoid ependymoma of the cerebral convexity.

A 29-year-old man had two episodes of seizures in 9 months. Following the first seizure he developed hemiparesis, which gradually progressed. Investigations revealed a large cerebral convexity mass lesion simulating a meningioma. During operation, a reddish-brown and well-defined tumor located in the subarachnoid space was encountered. Histopathological examination revealed a typical ependymoma. Extra-axial cerebral convexity ependymomas are extremely rare. The case is reported and a brief review of six similar cases reported in the literature is undertaken.

Spinal arteriovenous malformation associated with a radicular arteriovenous fistula suggested a metameric disease. A case report.

SUMMARY: A spinal intramedullary arteriovenous malformation (AVM) associated with a radicular arteriovenous fistula (AVF) is reported. The patient had mild myelopathy and low back pain. Spinal angiography revealed the AVM fed by the anterior spinal artery via left T10, T11 and right L1 radiculomedullary arteries and the radiculopial arteries of left L1, L2 and right T11, L3 levels and the radicular AVF at the left L4 level. There were three radiculomedullary arteries within four levels in our case. This spinal AVM associated with a radicular AVF is considered a genetic nonhereditary lesion with metameric link.

Simple decompression with small skin incision for cubital tunnel syndrome.

Eighteen elbows in 17 patients with cubital tunnel syndrome were treated by simple decompression using only a 1.5-2.5 cm skin incision with no endoscopic assistance. According to McGowan's criteria, three elbows were classified preoperatively as grade I, six as grade II and nine as grade III. The mean follow-up period was 14 months (range 3-25). Clinical results were evaluated as excellent for four elbows, good for ten and fair for four. Improvement of symptoms occurred in all patients and dislocation of the ulnar nerve was not observed. Simple decompression through a small skin incision can be recommended for the treatment of cubital tunnel syndrome, if the indication is appropriate.

A hypothesis of epiarachnoidal growth of vestibular schwannoma at the cerebello-pontine angle: surgical importance.

AIMS: The purpose of this study is to clarify the rearrangement of the arachnoid membrane on the vestibular schwannoma during its growth in relation to adjacent neurovascular structures for a better understanding of dissecting plane of arachnoid during surgery. METHODS: Arachnoid membrane over the tumour was investigated during surgery with suboccipital transmeatal approach in twenty-six tumours. All microsurgical procedures were recorded with a video and reviewed. The tumour growth was classified into five stages depending upon the tumour diameter in the cerebello-pontine (CP) angle: Stage 1; purely intracanalicular (2 cases), Stage 2; less than 5 mm (2 cases), Stage 3; > or = 5 and <15 mm (8 cases), Stage 4; > or = 15 and <25 mm (9 cases) and Stage 5; > or = 25 mm (5 cases). Rearrangement of the arachnoid on the tumour was conceptualised throughout all stages. RESULTS: All tumours of Stage 1 and 2 were entirely located in the subarachnoid space of the cerebello-pontine cistern without arachnoidal rearrangement, while all tumours of Stages 3 to 5 were enveloped, in the CP angle, with invaginated arachnoid membrane consisting of cerebello-pontine cistern except two surfaces; the medial pole and the tumour surface under the facial and cochlear nerves near the porus. CONCLUSION: The tumour originates subarachnoidally within the internal auditory meatus (IAM) and grows epiarachnoidally in the CP angle. Rearrangement of the arachnoid begins with its adhesion on the medial pole of the tumour along the porus, resulting in the arachnoidal invagination into the cerebello-pontine cistern with further growing of the tumour.

Development of lumbosacral spina bifida: three-dimensional computer graphic study of human embryos at Carnegie stage twelve.

There is still some controversy as to whether sacral spina bifida in humans is the result of a defect of the primary or secondary neural tube. As somites are related to the development of vertebrae and the primary neural tube is related to the development of the spinal cord in embryos, it is very important to determine the number of somites in normal human embryos at the time of closure of the primary neural tube to understand the contribution of primary neural tube defects to the development of spina bifida. However, in the literature, the number of somites in stage 12 human embryos is still controversial. The aim of this study is to find the number of somites in human embryos at Carnegie stage 12. Four human embryos at Carnegie stage 12 were selected from the laboratory of the Congenital Anomaly Research Center in Japan. The neural tube and somites were reconstructed from their slices by a three-dimensional computer graphic reconstruction technique. The reconstructed embryos were examined from multidirectional magnified images. Thirty-three pairs of somites were present in all these reconstructed embryos. As the 33rd pair of somites corresponds to the fifth sacral segment, the presence of 33 pairs of somites at Carnegie stage 12 suggests that spina bifida develops from defects of the primary neural tube.