Effect of Diluents on Active Pharmaceutical Ingredient Loss Until Patients Ingest Powder Formulation.

In powder formulations, it is a problem that the required therapeutic dose is not obtained because of loss of the active pharmaceutical ingredient (API). In this study, we investigated three types of lactose diluents, which are widely used as pharmaceutical excipients, for dispensing prednisolone powder. Extra-fine crystalline lactose, commonly used as a diluent in compounding powder formulations, was used as a comparison. The effect of lactose on the API loss rate was examined by analyzing the amount of prednisolone in the powder formulation taken out of a single-dose package after dispensing. The results showed that Dilactose-F had the lowest API loss rate (22%), followed by powder lactose (37.8%), extra-fine crystalline lactose (45.9%), and crystal form lactose (48.6%), indicating that the use of Dilactose-F as a diluent significantly improved API loss when compounding the powder formulation. Because each mixture of commercial prednisolone powder and lactose was within acceptable uniformity and loss rate before packaging, we considered that API loss occurred when the powder was taken out of the single-dose package before patients ingested them. Then, the physical properties of these lactose types affecting the API loss rate were examined. Strong correlation was not found between flowability and the API loss rate, but particle size distribution and bulk density were strongly correlated with the API loss rate. Furthermore, Dilactose-F, which showed the lowest API loss rate, did not show an exothermic peak due to epimerization to anhydrous ß -lactose in differential scanning calorimetry and showed a peak specific to ß -lactose in powder X-ray diffractometer. These results suggested that in powder compounding where the API content is low, the physical properties of lactose, such as particle size distribution, bulk density, and crystalline form, are intricately related to API loss.

Effects of prior osteoporosis treatment on the treatment response of romosozumab followed by denosumab in patients with postmenopausal osteoporosis.

In patients with postmenopausal osteoporosis, prior osteoporosis treatment affected the bone mineral density increase of following treatment with 12 months of romosozumab, although it did not affect that of following treatment with 12 months of denosumab after romosozumab. PURPOSE: To investigate the effects of prior osteoporosis treatment on the response to treatment with romosozumab (ROMO) followed by denosumab (DMAb) in patients with postmenopausal osteoporosis. METHODS: In this prospective, observational, multicenter study, treatment-naïve patients (Naïve; n = 55) or patients previously treated with bisphosphonates (BP; n = 37), DMAb (DMAb; n = 45) or teriparatide (TPTD; n = 17) (mean age, 74.6 years; T-scores of the lumbar spine [LS] - 3.2 and total hip [TH] - 2.6) were switched to ROMO for 12 months, followed by DMAb for 12 months. Bone mineral density (BMD) and serum bone turnover markers were evaluated for 24 months. RESULTS: A BMD increase was observed at 12 and 24 months in the following patients: Naïve (18.2% and 22.0%), BP (10.2% and 12.1%), DMAb (6.6% and 9.7%), and TPTD (10.8% and 15.0%) (P < 0.001 between the groups at both 12 and 24 months) in LS and Naïve (5.5% and 8.3%), BP (2.9% and 4.1%), DMAb (0.6% and 2.2%), and TPTD (4.3% and 5.4%) (P < 0.01 between the groups at 12 months and P < 0.001 at 24 months) in TH, respectively. The BMD increase in LS from 12 to 24 months was negatively associated with the levels of bone resorption marker at 24 months. Incidences of major fragility fractures for the respective groups were as follows: Naïve (5.5%), BP (16.2%), DMAb (11.1%), and TPTD (5.9%). CONCLUSIONS: Previous treatment affected the BMD increase of following treatment with ROMO, although it did not affect that of following treatment with DMAb after ROMO.

Factors that predict in-hospital mortality in eosinophilic granulomatosis with polyangiitis.

BACKGROUND: Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare systemic small-vessel vasculitis associated with asthma, eosinophilia, and necrotizing vasculitis. EGPA is potentially life-threatening and often involves peripheral neuropathies, peptic ulcers, cerebral vessel disease, and cardiovascular disease. However, there is limited understanding of the prognostics factors for patients with EGPA. We investigated the clinical features and factors affecting patients' in-hospital mortality, using a national inpatient database in Japan. METHODS: We retrospectively collected data of EGPA patients who required hospitalization between July 2010 and March 2013, using the Diagnosis Procedure Combination database. We evaluated EGPA patients' characteristics and performed multivariate logistic regression analyses to assess the factors associated with in-hospital mortality. RESULTS: A total of 2195 EGPA patients were identified. The mean age was 61.9 years, 42.1% (924/2195) were male, and 41.6% (914/2195) had emergent admission. In-hospital deaths occurred in 97/2195 patients (4.4%). Higher in-hospital mortality was associated with age older than 65 years, disturbance of consciousness on admission, unscheduled admission, respiratory disease, cardio-cerebrovascular disease, renal disease, sepsis, and malignant disease on admission. Lower mortality was associated with female gender and peripheral neuropathies. CONCLUSIONS: Our study revealed the clinical features of EGPA patients who required hospitalization and the factors associated with their mortality. These results may be useful for physicians when assessing disease severity or treatments for hospitalized EGPA patients.

Stabilizing effect of total ankle arthroplasty by distal translation and lateralization of talus in varus ankle deformity.

BACKGROUND: In end-stage arthritis indicated for total ankle arthroplasty (TAA), full-thickness cartilage damage, subchondral bone defect/shaving, and fluttering of the talar dome occur, shortening the distance between the tibial and talar insertions of ligaments and leading to laxity of ligaments surrounding the ankle joint. Under such conditions, medial ligaments (including the deltoid ligament) would not be expected to function properly. To stabilize the ankle joint during the stance phase, medial ligament function under tension is important. This study therefore examined whether TAA contributes to lengthening of the medial tibio-talar joint as evaluated radiographically, as a preferable method for achieving tensile effects on medial ligaments. MATERIALS AND METHODS: Twenty-four feet with end-stage varus deformity of the ankle joint that underwent TAA were retrospectively investigated, excluding cases with any malleolar osteotomy or fracture. Distance between proximal and distal insertions of medial ligaments, lateralization of the talus, and talar tilt angle under valgus/varus stress condition were evaluated pre- and postoperatively. RESULTS: Distance between proximal and distal insertions of medial ligaments was significantly elongated after TAA. At the same time, the talus showed significant lateralization. Furthermore, talar tilt under valgus/varus stress conditions was also significantly reduced after TAA. CONCLUSION: TAA affects distal translation and lateralization of the talus in cases of varus ankle deformity. These effects might contribute to re-providing tensile force on lax medial ligaments, improving ligament function.

Nutritional profile of rodent diets impacts experimental reproducibility in microbiome preclinical research.

The lack of reproducibility of animal experimental results between laboratories, particularly in studies investigating the microbiota, has raised concern among the scientific community. Factors such as environment, stress and sex have been identified as contributors, whereas dietary composition has received less attention. This study firstly evaluated the use of commercially available rodent diets across research institutions, with 28 different diets reported by 45 survey respondents. Secondly, highly variable ingredient, FODMAP (Fermentable Oligo-, Di-, Mono-saccharides And Polyols) and gluten content was found between different commercially available rodent diets. Finally, 40 mice were randomized to four groups, each receiving a different commercially available rodent diet, and the dietary impact on cecal microbiota, short- and branched-chain fatty acid profiles was evaluated. The gut microbiota composition differed significantly between diets and sexes, with significantly different clusters in ß-diversity. Total BCFA were highest (p = 0.01) and SCFA were lowest (p = 0.03) in mice fed a diet lower in FODMAPs and gluten. These results suggest that nutritional composition of commercially available rodent diets impact gut microbiota profiles and fermentation patterns, with major implications for the reproducibility of results across laboratories. However, further studies are required to elucidate the specific dietary factors driving these changes.

Suppression of tumorigenicity, but not anchorage independence, of human cancer cells by new candidate tumor suppressor gene CapG.

Previously, we isolated a series of cell lines from a human diploid fibroblast lineage as a model for multistep tumorigenesis in humans. After passaging a single LT-transfected fibroblast clone, differently progressed cell lines were obtained, including immortalized, anchorage-independent and tumorigenic cell lines. In the present paper, we analysed the gene expression profiles of these model cell lines, and observed that expression of the CapG protein was lost in the tumorigenic cell line. To examine the possibility that loss of CapG protein expression was required for tumorigenic progression, we transfected CapG cDNA into the tumorigenic cell line and tested for tumor-forming ability in nude mice. Results showed that ectopic expression of CapG suppressed tumorigenicity, but not growth in soft agar or liquid medium. We also found that certain cancer cell lines including stomach cancer, lung cancer and melanoma had also lost CapG expression. One such cancer cell line AZ521 also became non-tumorigenic after the introduction of CapG cDNA. Moreover, we showed that CapG expression was repressed in small-cell lung cancer tissues. Together, our findings indicated that CapG is a new tumor suppressor gene involved in the tumorigenic progression of certain cancers.

[Extrapleural pneumonectomy for relapsed solitary fibrous tumors of the pleura with pleural dissemination].

We report a case of a 62-year-old female with a prior thoracotomy for solitary fibrous tumor of the diaphragmatic pleura. There was no clear evidence of malignant solitary fibrous tumor of the pleura (SFTP). In the 19th postoperative month, she had a disseminated recurrence of SFTP in the left thoracic cavity. There was no evidence of metastasis from medical imaging. Accordingly, a left extrapleural pneumonectomy was performed. Pathological examination revealed a disseminated recurrence of malignant SFTP, showing a higher grade of malignancy, because the resected specimen was identical to the only section suspicious of malignancy in the previous tumor. She had no complaint and kept better performance status until the 7th postoperative month after the re-resection, when she had a recurrence in the left thoracic cavity and dissemination in the peritoneal cavity. She died of the recurrence 15 months after the re-resection and 34 months after the prior thoracotomy.

Biallelic inactivation of the APC gene in hepatoblastoma.

Familial adenomatous polyposis (FAP) is an inherited disorder caused by germline mutation of the adenomatous polyposis coli (APC) gene. Increased risk of hepatoblastoma (HBL) in FAP kindreds has been reported. To determine whether inactivation of the APC gene plays a role in development of HBL, 13 sporadic infantile hepatic tumors were analyzed for genetic alterations in the APC gene. A PCR-mediated RNase protection analysis was performed to detect subtle genetic alterations in the mutation cluster region and in exons 3 and 4 of the APC gene. The results showed that a G to T transversion at the splice acceptor site of the intron 3-exon 4 junction had occurred in one HBL. Sequence analysis of normal tissue of the patient proved the mutation to be germinal. Southern blot analysis at the APC locus revealed that the tumor had lost the opposite allele and was isodisomic at this locus. RNA analysis indicated that the tumor contained only the small APC transcript, from which exon 4 was entirely absent. Since abnormal splicing causes termination due to frameshift, it was hypothesized that only the truncated APC protein was expressed in this tumor. These findings suggest that inactivation of the APC gene is closely related to tumorigenesis of HBLs in FAP patients.

A frequent alteration of p53 gene in carcinoma in adenoma of colon.

In general, colorectal carcinoma is thought to originate mainly from adenoma, and this pathway is called the adenoma-carcinoma sequence. Carcinoma in adenoma is an appropriate model for analysis of this mechanism, because adenoma and carcinoma tissues coexist in the same polyp and the carcinoma is thought to have originated from the surrounding adenoma. Expression of the p53 protein was analyzed in 36 cases of carcinoma in adenoma in the colon by immunohistochemistry using an anti-human p53 monoclonal antibody (PAb1801). Alterations of the p53 gene were analyzed by the polymerase chain reaction for microanalysis of normal mucosa, adenoma, and carcinoma from histological slides. Mutations were assessed by the polymerase chain reaction-single strand conformation polymorphism analysis and identified by DNA sequencing in some cases. Loss of heterozygosity was studied by polymerase chain reaction-restriction fragment length polymorphism analysis. Positive staining for p53 was detected in three (8%) of 37 adenomas and 20 (53%) of 38 focal carcinomas. One (7%) of 15 adenomas with mild dysplasia, three (14%) of 22 adenomas with moderate dysplasia, and 16 (42%) of 38 focal carcinomas had a mutation in exon 5 through exon 8 of the p53 gene. As for allelic loss in the p53 gene locus, only one adenoma with moderate dysplasia had loss of heterozygosity, whereas six (40%) of 15 focal carcinomas had loss of heterozygosity. Of those tumors (3 of 37 adenomas and 20 of 38 focal carcinomas) that reacted with PAb1801, 78% (18 of 23) showed genetic alterations. Among 52 tumors which showed negative staining, five tumors had a p53 mutation and four of them were nonsense mutations. Putting all of these results together, 71% (24 of 34) of the cases underwent p53 gene and protein alterations during the conversion from adenoma to focal carcinoma. These data clearly indicate that genetic alterations of p53 are involved mainly in the malignant transformation from adenoma to focal carcinoma in colon carcinogenesis. In addition, some cases show heterogeneity of the p53 gene in carcinoma in adenoma of the colon. There may be other pathways than p53 responsible for malignant change in the colon.

Ancient allelism at the cytosolic chaperonin-alpha-encoding gene of the zebrafish.

The T-complex protein 1, TCP1, gene codes for the CCT-alpha subunit of the group II chaperonins. The gene was first described in the house mouse, in which it is closely linked to the T locus at a distance of approximately 11 cM from the Mhc. In the zebrafish, Danio rerio, in which the T homolog is linked to the class I Mhc loci, the TCP1 locus segregates independently of both the T and the Mhc loci. Despite its conservation between species, the zebrafish TCP1 locus is highly polymorphic. In a sample of 15 individuals and the screening of a cDNA library, 12 different alleles were found, and some of the allelic pairs were found to differ by up to nine nucleotides in a 275-bp-long stretch of sequence. The substitutions occur in both translated and untranslated regions, but in the former they occur predominantly at synonymous codon sites. Phylogenetically, the alleles fall into two groups distinguished also by the presence or absence of a 10-bp insertion/deletion in the 3' untranslated region. The two groups may have diverged as long as 3.5 mya, and the polymorphic differences may have accumulated by genetic drift in geographically isolated populations.

Body fatness and fat distribution as predictors of metabolic abnormalities and early carotid atherosclerosis.

OBJECTIVE: To test the hypothesis that intra-abdominal fat plays a primary role over general adiposity for metabolic abnormalities and atherosclerosis. RESEARCH DESIGN AND METHODS: We cross-sectionally studied 849 Japanese men aged 50.3 +/- 8.5 years (range 20-78) with BMI 23.5 +/- 2.9 kg/m(2). Intimal-medial thickness (IMT) of the carotid artery was measured by ultrasound. General adiposity was assessed by BMI. Waist circumference and waist-to-hip ratio (WHR) were used as a surrogate measure for abdominal fat. Abdominal subcutaneous fat area (ASF) and intra-abdominal fat area (IAF) were measured by computed tomography. Correlations between these measures and carotid IMT were analyzed. The interaction of generalized adiposity (BMI) and IAF in relation to metabolic variables, such as glucose tolerance, insulin resistance, and serum lipids, was also evaluated. RESULTS: BMI, waist circumference, WHR, ASF, and IAF were all correlated with carotid IMT. Adjustment for BMI eliminated the associations between IMT and waist circumference, ASF, and IAF. In contrast, WHR retained a significant correlation with IMT. BMI and IAF were associated with insulin resistance, glucose tolerance, HDL cholesterol, and blood pressure independently of each other. IAF was an independent correlate for serum triglyceride, but BMI was not. CONCLUSIONS: The primary importance of IAF over general adiposity for carotid atherosclerosis was not confirmed. Caution is recommended when using WHR as a measure of abdominal fat. The roles of IAF for metabolic abnormalities may be more limited than conventionally thought. BMI and WHR are simple and better clinical predictors for carotid atherosclerosis versus IAF.

Survey of Cryptosporidium spp. and Giardia spp. infections in various animals at a zoo in Japan.

A total of 284 fecal samples of 89 species (43 mammalian species and 46 avian species) were examined for Cryptosporidium oocysts and Giardia cysts from 1999 to 2002. Each sample was collected at the zoo located at Osaka in Japan and examined by microscopy after performing the sucrose flotation method and by two immunofluorescent assay kits for detection of Cryptosporidium oocysts and Giardia cysts. Cryptosporidium spp. was found only in a raccoon dog (Nyctereutes procyonoides), and Giardia spp. was detected in a mandarin duck (Aix galericulata) and two ruddy shelducks (Tadorna ferruginea). In this study, the prevalences of these parasites were found to be low. However, these results suggested that the infected animals could serve as a source of contamination for surface water. This is the first report about the survey of Cryptosporidium spp. and Giardia spp. at a zoo in Japan.

Association between serum leptin concentrations and bone mineral density, and biochemical markers of bone turnover in adult men.

Leptin, the product of the ob gene, has been shown to inhibit bone formation in mice. We addressed whether leptin has any role in the regulation of bone mineral density (BMD) in humans. Subjects were 221 adult men with a mean (+/-SD) age and body mass index of 52.1 +/- 8.7 yr and 23.6 +/- 2.8 kg/m2. Serum leptin, carboxyterminal propeptide of type 1 procollagen (PICP; a marker of bone formation), and cross-linked carboxyterminal teleopeptide of type 1 collagen (a marker of bone resorption) were measured by RIA. BMD was assessed by single photon absorptiometry, and total fat mass was determined by bioimpedance analysis. BMD was inversely associated with serum leptin concentrations and total fat mass after adjustment for body weight. PICP, but not cross-linked carboxyterminal teleopeptide of type 1 collagen, was inversely correlated with serum leptin. These results may suggest that an increase in serum leptin reduces bone formation and decreases BMD in adult men. Leptin may be a regulator of BMD in humans.

Functional characterization of the rat multispecific organic anion transporter OAT1 mediating basolateral uptake of anionic drugs in the kidney.

The functional characteristics of rat organic anion transporter OAT1 were investigated using Xenopus laevis oocytes. Uptake of p-aminohippurate (PAH) by the oocytes expressing OAT1 was markedly inhibited by glutarate, alpha-ketoglutarate and probenecid, moderately inhibited by folate and methotrexate, but not inhibited by taurocholate or tetraethylammonium. Methotrexate and folate were transported by OAT1, but probenecid, a typical inhibitor of organic anion transporter, was not transported. Inhibition of PAH uptake by aliphatic dicarboxylates with various alkyl chain lengths was maximal at 5 (glutarate) and 6 (adipate) carbon atoms. OAT1-mediated PAH uptake was markedly inhibited by phorbol 12-myristate 13-acetate (PMA), phorbol 12,13-dibutyrate and mezerein, but not by 4alpha-phorbol 12,13-didecanoate. The inhibitory effect of PMA was attenuated in the presence of staurosporine, suggesting that OAT1 is regulated by protein kinase C. These results suggest that the substrate recognition of OAT1 is comparable to that of renal basolateral organic anion transporter, and the transport activity is regulated by protein kinase C.

Transmembrane TNF (pro-TNF) is palmitoylated.

Human transmembrane tumor necrosis factor (pro-TNF) was examined for protein acylation. The cDNA encoding pro-TNF was expressed in both COS-1 cells and Sf9 cells and metabolic labeling with [(3)H]myristic or [(3)H]palmitic acid was attempted. The 17 kDa mature TNF secreted from the transfected cells was not labeled, whereas the 26 kDa pro-TNF was specifically labeled with [(3)H]palmitic acid. The [(3)H]palmitic acid labeling of pro-TNF was eliminated by treatment with hydroxylamine, indicating that the labeling was due to palmitoylation of a cysteine residue via a thioester bond. Site-directed mutagenesis of the two cysteine residues residing in the leader sequence of pro-TNF demonstrated that palmitoylation of pro-TNF occurs solely at Cys-47, located at the boundary between the transmembrane and cytoplasmic domains of pro-TNF. Thus, pro-TNF interacts with the plasma membrane via both its proteinaceous transmembrane domain and a lipid anchor.

p53-regulated GML gene expression in non-small cell lung cancer. a promising relationship to cisplatin chemosensitivity.

The GML gene (glycosylphosphatidylinositol-anchored molecule-like protein gene) is a novel gene specifically induced by wild-type p53, which may participate in cell cycle control or the cell apoptotic pathway. Recent experiments suggest that the expression of this novel gene in cancer cells is closely associated with sensitivity to certain anticancer drugs. To elucidate the role of the gene expression in cisplatin (CDDP) chemosensitivity of non-small cell lung cancer (NSCLC), 30 surgically resected materials were examined by reverse transcriptase-polymerase chain reaction (RT-PCR). GML gene expression was detected in 9 (30%) samples. Its incidence was significantly higher in immunohistochemically p53-negative (P=0.040) or wild-type p53 tissues (P=0.041). On in vitro chemosensitivity testing using 29 primary tissues, six samples with GML gene expression showed good sensitivity to CDDP. In particular, in tissues with immunohistochemically p53-negative accumulation, those with GML gene expression showed significantly better in vitro sensitivity to CDDP (P=0.012). Clinically a good response to CDDP-based chemo(thermo)therapy for NSCLC patients with tumour residue or recurrence, was observed only in those with p53-negative accumulation and GML gene expression, in agreement with in vitro results. Thus, although the number of tested samples was small, GML gene expression is commonly detected in immunohistochemically p53-negative NSCLCs in close association with good sensitivity to CDDP. GML gene expression analysis may serve as a predictor of CDDP-based chemotherapy for patients with NSCLC.

Localization of chick retinal visinin-like immunoreactivity in the rat forebrain and diencephalon.

The present study is an examination, using an indirect immunofluorescence method, of the distribution of visinin, a 24,000 dalton peptide, in the rat forebrain and diencephalon. Immunoreactive structures were localized in the neuronal elements showing an uneven distribution. Immunoreactive neurons were found in the olfactory bulb, anterior olfactory nucleus, cerebral cortex, amygdaloid complex, ventral portion of the nucleus caudatus putamen, septal area, nucleus accumbens, nucleus paratenialis, nucleus rhomboideus, nucleus reuniens, nucleus paraventricularis hypothalami, nucleus supraopticus, nucleus anterior hypothalami, preoptic area, hypothalamic periventricular nucleus, nucleus mammillaris medialis, medial habenular nucleus, zona incerta, nucleus lateralis thalami, nucleus tractus optici and gyrus dentatus. Immunoreactive fibers were observed in the above areas, particularly near the labelled cells, forming fiber plexuses of varying density. In addition, dense plexuses were also seen in the globus pallidus, anteroventral nucleus of the thalamus, substantia nigra and hippocampus. In the former three structures, no labelled cells were present and in the latter, a few scattered neurons were found, indicating that these fibers originate from extrinsic sources.

Localization of chick retinal 24,000 dalton protein (visinin)-like immunoreactivity in the rat lower brain stem.

The distribution of visinin, a 24,000 dalton peptide, in the lower brain stem of the rat was examined by means of an indirect immunofluorescent method. Visinin-immunoreactive structures were found to be unevenly distributed only in the neuronal elements. The following neuronal systems were strongly labeled by the antiserum; the Purkinje cell system, mammillotegmental system, habenulointerpeduncular system, the second layer of the superior colliculus, ventral tegmental area, substantia nigra pars lateralis, area medial to the medial geniculate body, parabrachial area, dorsal and ventral nuclei of the lateral lemniscus, pontine reticular formation just medial to the trigeminal principal nucleus, superior olivary nucleus, solitarii nucleus, external layer of the inferior colliculus and spinal trigeminal nucleus. The densities of the labeled fibers in these areas paralleled those of the labeled cells. In addition, highly dense visinin-immunoreactive fiber plexuses were seen in the zona compacta of the substantia nigra, lateral portion of the interpeduncular nucleus, ventral tegmental nucleus of Gudden and vestibular nucleus.

Analysis of neurotrophin-3 expression using the lacZ reporter gene suggests its local mode of neurotrophic activity.

We replaced the mouse neurotrophin-3 gene with the Escherichia coli-derived lacZ gene by means of homologous recombination. The mice with this mutation were useful models for studying the distribution of neurotrophin-3 expression in vivo, because visualization by 5-bromo-4-chloro-3-indoyl-beta-D-galactopyranoside (X-Gal) staining was simple and rapid compared with in situ hybridization or immunohistochemistry. Whole-mount staining of mutant embryos at embryonic day 10 revealed that lacZ, a reporter for the neurotrophin-3 gene, was expressed in the mesencephalon, mandibular arch and somites. In the embryos at days 13-17, lacZ was markedly expressed in the peripheral target tissues of sensory and sympathetic neurons. We also found that spinal motor neurons and sensory neurons in trigeminal and dorsal root ganglia express lacZ. Some of these X-Gal staining regions overlapped with the sites expressing trkC, a high-affinity receptor for neurotrophin-3. The distribution of X-Gal staining in heterozygotes and homozygotes was similar to that of neurotrophin-3 messenger RNA detected by in situ hybridization. However, there was less lacZ expression in the dorsal root ganglia of homozygotes than neurotrophin-3 expression in wild-type mice. These results suggest that the neurotrophin-3 produced in the dorsal root ganglia also plays a role in the survival of some of the neurotrophin-3-positive neurons and that the local mode of neurotrophic activity is widely distributed.

Distribution of calcitonin gene-related peptide in the rat peripheral nervous system with reference to its coexistence with substance P.

This immunocytochemical study, using a double-staining method, showed that calcitonin gene-related peptide-like immunoreactive structures are widely distributed in the peripheral nervous system and that many of them coexist with substance P-like immunoreactive structures in single sensory ganglion cells. Neurons positive for calcitonin gene-related peptide but negative for substance P were detected in sensory ganglia. These cells were large (about 30-45 micron in diameter); these primary sensory neurons containing calcitonin gene-related peptide can probably act independently of substance P. There were neurons containing calcitonin gene-related peptide without substance P in the pterygopalatine ganglion, although these cells were less numerous than in the sensory ganglia. In consecutive sections, calcitonin gene-related peptide-like structures occurred in thyroid parafollicular cells, which also contain calcitonin. This suggested that messenger RNA for producing calcitonin gene-related peptide is also present in the thyroid, and like calcitonin, calcitonin gene-related peptide may have a peripheral physiological role.