Identification of AFAP1L1 as a prognostic marker for spindle cell sarcomas.

Spindle cell sarcomas consist of tumors with different biological features, of which distant metastasis is the most ominous sign for a poor prognosis. However, metastasis is difficult to predict on the basis of current histopathological analyses. We have identified actin filament-associated protein 1-like 1 (AFAP1L1) as a candidate for a metastasis-predicting marker from the gene expression profiles of 65 spindle cell sarcomas. A multivariate analysis determined that AFAP1L1 was an independent factor for predicting the occurrence of distant metastasis (P=0.0001), which was further confirmed in another set of 41 tumors by a quantitative mRNA expression analysis. Immunohistochemical staining using paraffin-embedded tumor tissues revealed that the metastasis-free rate was significantly better in tumors negative for AFAP1L1 (P=0.0093 by log-rank test). Knocking down the AFAP1L1 gene in sarcoma cells resulted in inhibition of the cell invasion, and forced expression of AFAP1L1 in immortalized human mesenchymal stem cells induced anchorage-independent growth and increased cell invasiveness with high activity levels of matrix metallopeptidase. Furthermore, tumor growth in vivo was accelerated in AFAP1L1-transduced sarcoma cell lines. These results suggest that AFAP1L1 has a role in the progression of spindle cell sarcomas and is a prognostic biomarker.

Mediastinal synovial sarcoma.

Synovial sarcoma of the mediastinum is a rare neoplasm that has overlapping histological and immunophenotypic features with other tumors in the differential diagnosis. We describe a case of this disease. The tumor was located at the right side of the pericardium, where an FDG-PET scan showed an uptake. It was resected, a resection which was complicated by the necessity of partially resecting the pericaridium and right middle lobe which were invaded by the tumor. The doubling time of the main tumor was 11.8 days. The margin of the resected specimen was tumor-free both macro- and microscopically. Reverse transcription-PCR confirmed the diagnosis of synovial sarcoma. The patient rejected chemotherapy or radiation therapy, and had recurrent tumors only one month after the operation. Finally, she opted to have only palliative care and died 79 days after the operation.

PDGFR-A is a therapeutic target in alveolar rhabdomyosarcoma.

Alveolar rhabdomyosarcoma is an aggressive skeletal muscle cancer of childhood. Our initial studies of rhabdomyosarcoma gene expression for patients enrolled in a national clinical trial suggested that platelet-derived growth factor receptor A (PDGFR-A) may be a mediator of disease progression and metastasis. Using our conditional mouse tumor models that authentically recapitulate the primary mutations and metastatic progression of alveolar rhabdomyosarcomas in humans, we found by immunoblotting and immunokinase assays that PDGFR-A and its downstream effectors, mitogen-activated protein kinase and Akt, were highly activated in both primary and metastatic tumors. Inhibition of PDGFR-A by RNA interference, small molecule inhibitor or neutralizing antibody had a dramatic effect on tumor cell growth both in vitro and in vivo, although resistance evolved in one-third of tumors. These results establish proof-of-principal for PDGFR-A as a therapeutic target in alveolar rhabdomyosarcoma.

Expression of the cadherin-11 gene is a discriminative factor between articular and growth plate chondrocytes.

OBJECTIVE: Calcification of hypertrophic chondrocytes is the final step in the differentiation of growth plates, although the precise mechanism is not known. We have established two growth plate-derived chondrocyte cell lines, MMR14 and MMR17, from p53-/- mice (Nakamata T, Aoyama T, Okamoto T, Hosaka T, Nishijo K, Nakayama T, et al. In vitro demonstration of cell-to-cell interaction in growth plate cartilage using chondrocytes established from p53-/- mice. J Bone Miner Res 2003;18:97-107). Prolonged in vitro culture produced calcified nodules in MMR14, but not in MMR17. Factors responsible for the difference in calcification between the two cell lines may also be involved in the physiological calcification in growth plate. DESIGN: Gene expression profiles of MMR14 and MMR17 were compared using a cDNA microarray to identify candidate genes involved in the calcification process. RESULTS: Forty-five genes were identified as upregulated in MMR14, including the cadherin-11 (Cdh-11) gene. The expression of Cdh-11 in MMR14 was detected in cell-cell junctions, while no expression was observed in MMR17. Primary cultured chondrocytes from growth plate (GC) also expressed the Cdh-11, and the staining of Cdh-11 was observed in the late hypertrophic zone of growth plate. Cell aggregation assays showed that chondrocytes required Ca2+ to form nodules, and knockdown of the Cdh-11 gene expression using short interfering RNA inhibited the formation of calcified nodules in MMR14. The introduction of Cdh-11 into MMR17 failed to produce calcified nodules indicating that Cdh-11 is one, but not the sole, factor responsible for the production of calcified nodules. CONCLUSION: Although the physiological role is still unclear, Cdh-11 is a discriminative factor between articular and growth plate chondrocytes.

Electro-acupuncture stimulation to a hindpaw and a hind leg produces different reflex responses in sympathoadrenal medullary function in anesthetized rats.

The effects of electro-acupuncture stimulation (EAS) of two different areas of a hindlimb with different stimulus intensities on sympathoadrenal medullary functions were examined in anesthetized artificially ventilated rats. Two needles of 160 microm diameter and about 5 mm apart were inserted about 5 mm deep into a hindpaw (Chungyang, S42) or a hind leg (Tsusanli, S36) and current of various intensities passed to excite various afferent nerve fiber groups at a repetition rate of 20 Hz and pulse duration of 0.5 ms for 30-60 s. Fiber groups of afferent nerves stimulated in a hindlimb were monitored by recording evoked action potentials from the afferents innervating the areas stimulated. The sympathoadrenal medullary functions were monitored by recording adrenal sympathetic efferent nerve activity and secretion rates of catecholamines from the adrenal medulla. EAS of a hindpaw at a stimulus strength sufficient to excite the group III and IV somatic afferent fibers produced reflex increases in both adrenal sympathetic efferent nerve activity and the secretion rate of catecholamines. EAS of a hind leg at a stimulus strength sufficient to excite the group III and IV afferent fibers produced reflex responses of either increases or decreases in sympathoadrenal medullary functions. All responses of adrenal sympathetic efferent nerve activity were lost after cutting the afferent nerves ipsilateral to the stimulated areas, indicating that the responses are the reflexes whose afferents nerve pathway is composed of hindlimb somatic nerves. It is concluded that electro-acupuncture stimulation of a hindpaw causes an excitatory reflex, while that of a hind leg causes either excitatory or inhibitory reflex of sympathoadrenal medullary functions, even if both group III and IV somatic afferent fibers are stimulated.

Adverse events in acupuncture and moxibustion treatment: a six-year survey at a national clinic in Japan.

OBJECTIVES: Many of the frequently reported adverse effects of acupuncture are serious or severe such as pneumothorax, infection, spinal cord injury, or cardiac injury. However, reviewing published case reports does not provide enough information to assess the safety of acupuncture and moxibustion. In order to investigate adverse events of acupuncture, we reviewed all the relevant cases reported by the therapists at our clinic. SETTING: Over a 6 year period, in the national Tsukuba College of Technology Clinic in Japan, all the acupuncture and moxibustion therapists were required to report the cases of adverse events immediately upon recognition. RESULTS: A total of 84 therapists (13 preceptors and 71 interns) participated in the treatments. The total number of treatments was 65,482. Ninety four (0.14%) adverse events were reported. There were fourteen categories: failure to remove needles (27 cases), ecchymosis or hematoma without pain (9 cases), ecchymosis or hematoma accompanied by pain (8 cases), burn injury (7 cases), discomfort (7 cases), dizziness (6 cases), nausea or vomiting (6 cases), pain in the punctured region (6 cases), minor hemorrhage (4 cases), aggravation of complaint (4 cases), malaise (3 cases), suspected contact dermatitis (3 cases), fever (3 cases) and numbness in the upper extremities (1 case). CONCLUSION: No serious or severe cases such as pneumothorax, infection, or spinal cord injury were reported by the college preceptors and interns. The results indicate that serious or severe adverse events are rare in standard practice. We suggest that most severe or serious cases of adverse events caused by acupuncture reported in journals are actually cases of negligence. In the future, negligence should be discussed from the point of view of medical education and technical instruction for the therapists, and adverse reactions should be discussed from the point of view of incidence and prevention based on the result of further investigation.

Dynamic electromyographic response following acupuncture: possible influence on synergistic coordination.

Thirty healthy subject's left and right lumber paraspinal (LP) EMG activity was recorded during a trunk flexion-return movement and the maximum integrated EMG amplitude (absolute EMG) during this movement in each side was compared. Twenty subjects showed less than 20% difference between the left and right side (symmetrical subjects-SS) and 10 subjects showed more than a 20% difference (asymmetrical subjects AS). As were administered acupuncture stimulation on LP muscles. Significant reduction in lumbar EMG asymmetry was observed after acupuncture stimulation (exact p=.049). No specific pattern of response in absolute EMG values was observed in the stimulate side. On the nonstimulated side, there was a significant reduction in absolute EMG values when the baseline value for that side was high (p=.037) and a significant increase when it was low (p=.0185). The results suggest that acupuncture may be beneficial for decreasing functional muscular distortion and improving synergistic coordination.

Effects of moxibustion on the enhancement of serum antibody in rabbit against Staphylococcus aureus.

The titer and activity of antibody in rabbits immunized with heat-killed vaccine were assessed with and without moxibustion treatment. Enzyme-linked immunosorbent assay (ELISA) was applied for the detection of immunoglobulins to Smith strain of Staphylococcus aureus. Positive IgM titer of more than 0.4 were observed with this assay against the moxibustion group (P < 0.05). The titer of IgG antibody also increased; however, there was no significant difference between the moxibustion group and the control group. The ELISA inhibition test showed significantly higher protective activity of the sera in the moxibustion group at the 9th week after the first immunization (P < 0.05).

The physiological responses induced by superficial acupuncture: a comparative study of acupuncture stimulation during exhalation phase and continuous stimulation.

This study investigated the physiological effect of superficial acupuncture stimulation during a patient's exhalation phase in a sitting position (SES). The response to SES was compared to the stimulation applied continuously without considering the respiratory phase (CONT). It evaluated a chronic tension-type headache patient's static electromyographic (EMG) activity, pain response, heart rate, pulse height, and skin conductance level. The results indicated that SES stimulation significantly decreased headache intensity and demonstrated a strong trend towards decreasing static EMG activity compared to CONT stimulation. The study concluded that acupuncture, applied on the same point and at the same depth, produced different physiological effects, depending on whether the stimulation was applied during exhalation only or continuously applied. This suggests that the effect of acupuncture derives not only from point selection matching symptoms, but also from a consideration and utilization of the patient's respiratory phase during stimulation.

Decreased heart rate by acupuncture stimulation in humans via facilitation of cardiac vagal activity and suppression of cardiac sympathetic nerve.

The effect of acupuncture stimulation applied to a Ximen point (P4) of a forearm on heart rate was studied in healthy volunteer human subjects. Acupuncture stimulation decreased heart rate, or gave no significant response. The decreased response of heart rate following acupuncture was attenuated by administration of atropine and propranolol. Therefore, the acupuncture-induced response of decrease in heart rate was concluded to be a result of a reciprocal coordination of an increase in cardiac vagal activity and a decrease in cardiac sympathetic activity.

DAX-1 gene mutations and deletions in Japanese patients with adrenal hypoplasia congenita and hypogonadotropic hypogonadism.

Abnormality of the DAX-1 gene accounts for many instances of congenital adrenal hypoplasia. In the present study, we performed molecular genetic analysis of DAX-1 in 4 unrelated Japanese patients with adrenal hypoplasia congenita and hypogonadotropic hypogonadism. A double-point mutation for V126M and W171X was identified in 1 family and a complex de novo insertion-deletion mutation was identified in a second. The DAX-1 gene was entirely deleted in a 3rd patient as well as in a 4th with the additional feature of glycerol kinase deficiency.

Study on the primary visual cortex of visually impaired subjects by means of 123I-IMP SPECT and MRI.

We conducted a study of rCBF in the primary visual cortex of visually impaired subjects who have not been subjected to external stimulation for a long period, by means of 123I-IMP SPECT and MRI. The four subjects had lost their sight due to brain tumors (n = 2), glaucoma (n = 1) and trauma (n = 1). 123I-IMP SPECT showed no differences between the visually impaired group and a visually sound control group on visual analysis as well as semiquantitative analysis. MRI of the visually impaired subjects showed no organic changes, such as atrophy, in the occipital cortex. In conclusion, visually impaired subjects have no decrease in rCBF and no anatomical changes in the primary visual cortex.

Neural mechanism of depressor responses of arterial pressure elicited by acupuncture-like stimulation to a hindlimb in anesthetized rats.

The effects of acupuncture-like stimulation of a hindlimb on renal sympathetic nerve activity (RNA) as well as mean arterial blood pressure (MAP) were examined in anesthetized rats. An acupuncture needle (diameter of 160 microns) was inserted into the skin of a hindlimb and underlying muscles to a depth of 5 mm and was twisted at about 1 Hz. Under deep anesthetic condition, in about 70% of trials, acupuncture-like stimulation for 60 s induced a decrease in MAP which was accompanied by a decrease in RNA. Acupuncture-like stimulation applied to the muscles alone, but not to the skin alone, induced inhibition of RNA and MAP. Transection of sciatic and femoral nerves ipsilateral to the hindlimb stimulation completely abolished the responses of RNA and MAP. The hindlimb stimulation excited the femoral and common peroneal afferent nerves. In spinalized animals, the hindlimb stimulation did not produce any changes in RNA and MAP. The results indicate that the decrease in MAP induced by acupuncture-like stimulation of a hindlimb is a reflex response. The afferent pathway is composed of hindlimb muscle afferents while the efferent pathway is composed of sympathetic vasoconstrictors including the renal nerves. Endogenous opioids may not be involved in the present reflex, because an intravenous injection of naloxone, an antagonist of the opioid receptors, did not influence the reflex.

Stimulation of lumbar sympathetic trunk produces vasoconstriction of the vasa nervorum in the sciatic nerve via alpha-adrenergic receptors in rats.

The effects of repetitive electrical stimulation of a lumbar sympathetic trunk (LST) for 30 s at various frequencies and supramaximum intensity on the nerve blood flow in a sciatic nerve were studied by laser Doppler flowmetry in anesthetized Fischer-344 male rats. The response was biphasic; i.e. an initial increase and then a decrease. The maximum mean increase after 2 Hz stimulation was 22 +/- 8%, while the maximum mean decrease after 20-50 Hz stimulation was 79 +/- 3%, of the prestimulus control level. The initial increase, which was greater at lower frequencies and existed even after local sympathetic denervation, was passive, and was caused by the systemic pressor response to LST stimulation. The decrease, which was nearly abolished by an i.v. alpha-adrenergic blocker, phentolamine (10 mg/kg), resulted from vasoconstriction in the vasa nervorum, mainly via activation of alpha-adrenergic receptors.

Deoxyribonucleic acid analyses of five families with familial inherited thyroid stimulating hormone deficiency.

Five families with familial inherited TSH deficiency, reported to date, were examined for the TSH beta gene at the nucleotide level. The first family carries a single base substitution in the 29th codon which lies in the so-called CAGYC region; GCA (glycine) is replaced by AGA (arginine). This substitution induces conformational changes of the beta-polypeptide which make it unable to associate with the alpha-subunit. This mutation generates a new cleavage site for a restriction endonuclease MaeI, a new marker that can be used for DNA diagnosis. The second and third families were found to carry the same nucleotide substitution. Also, all three families were associated with an additional single base substitution in intron 2 as a polymorphic change, suggesting that these three families may have originated from the same single founder from Shikoku Island in Japan. The nucleotide sequence from the fourth and fifth families showed no alterations in the TSH beta gene from the about -200 basepair up-stream region to the polyadenylation site.