The effect of anastomotic leakage on the incidence of recurrence after tri-modality therapy for esophageal adenocarcinomas.

BACKGROUND: Neoadjuvant chemoradiotherapy (nCRTx) reduces the incidence of recurrence, while anastomotic leakage has shown increase the risk of recurrence. The primary objective of this retrospective study was to investigate the incidence and pattern of recurrence and secondary median recurrence-free interval and post-recurrence survival in patients with and without anastomotic leakage after multimodal therapy for esophageal adenocarcinoma. METHODS: Patients with recurrence after multimodal therapy between 2010 and 2018 were included. RESULTS: Six hundred and eighteen patients were included, 91 (14.7%) had leakage and 278 (45.0%) recurrence. Patients with leakage did not develop recurrence more often (48.4%) than those without (44.4%, [p = 0.484]). Recurrence-free interval for patients with (n = 44) and without leakage (n = 234) was 39 and 52 weeks, respectively (p = 0.049). Post-recurrence survival was 11 and 16 weeks, respectively (p = 0.702). Specified by recurrence site, post-recurrence survival for loco-regional recurrences was 27 versus 33 weeks (p = 0.387) for patients with and without leakage, for distant 9 versus 13 (p = 0.999), and for combined 11 versus 18 weeks (p = 0.492). CONCLUSION AND DISCUSSION: No higher incidence of recurrent disease was observed in patients with anastomotic leakage, however it is associated with a shorter recurrence-free interval. This could have implications for surveillance, as early detection of recurrent disease could influence therapeutic options.

Machine learning models in clinical practice for the prediction of postoperative complications after major abdominal surgery.

Complications after surgery have a major impact on short- and long-term outcomes, and decades of technological advancement have not yet led to the eradication of their risk. The accurate prediction of complications, recently enhanced by the development of machine learning algorithms, has the potential to completely reshape surgical patient management. In this paper, we reflect on multiple issues facing the implementation of machine learning, from the development to the actual implementation of machine learning models in daily clinical practice, providing suggestions on the use of machine learning models for predicting postoperative complications after major abdominal surgery.

Short-term outcome for high-risk patients after esophagectomy.

Transthoracic esophagectomy (TTE) for esophageal cancer facilitates mediastinal dissection; however, it has a significant impact on cardiopulmonary status. High-risk patients may therefore be better candidates for transhiatal esophagectomy (THE) in order to prevent serious complications. This study addressed short-term outcome following TTE and THE in patients that are considered to have a higher risk of surgery-related morbidity. This population-based study included patients who underwent a curative esophagectomy between 2011 and 2018, registered in the Dutch Upper GI Cancer Audit. The Charlson comorbidity index was used to assign patients to a low-risk (score ≤ 1) and high-risk group (score ≥ 2). Propensity score matching was applied to produce comparable groups between high-risk patients receiving TTE and THE. Primary endpoint was mortality (in-hospital/30-day mortality), secondary endpoints included morbidity and oncological outcomes. Additionally, a matched subgroup analysis was performed, including only cervical reconstructions. Of 5,438 patients, 945 and 431 high-risk patients underwent TTE and THE, respectively. After propensity score matching, mortality (6.3 vs 3.3%, P = 0.050), overall morbidity, Clavien-Dindo ≥ 3 complications, pulmonary complications, cardiac complications and re-interventions were significantly more observed after TTE compared to THE. A significantly higher mortality after TTE with a cervical reconstruction was found compared to THE (7.0 vs. 2.2%, P = 0.020). Patients with a high Charlson comorbidity index predispose for a complicated postoperative course after esophagectomy, this was more outspoken after TTE compared to THE. In daily practice, these outcomes should be balanced with the lower lymph node yield, but comparable positive node count and radicality after THE.

Machine learning versus logistic regression for the prediction of complications after pancreatoduodenectomy.

BACKGROUND: Machine learning is increasingly advocated to develop prediction models for postoperative complications. It is, however, unclear if machine learning is superior to logistic regression when using structured clinical data. Postoperative pancreatic fistula and delayed gastric emptying are the two most common complications with the biggest impact on patient condition and length of hospital stay after pancreatoduodenectomy. This study aimed to compare the performance of machine learning and logistic regression in predicting pancreatic fistula and delayed gastric emptying after pancreatoduodenectomy. METHODS: This retrospective observational study used nationwide data from 16 centers in the Dutch Pancreatic Cancer Audit between January 2014 and January 2021. The area under the curve of a machine learning and logistic regression model for clinically relevant postoperative pancreatic fistula and delayed gastric emptying were compared. RESULTS: Overall, 799 (16.3%) patients developed a postoperative pancreatic fistula, and 943 developed (19.2%) delayed gastric emptying. For postoperative pancreatic fistula, the area under the curve of the machine learning model was 0.74, and the area under the curve of the logistic regression model was 0.73. For delayed gastric emptying, the area under the curve of the machine learning model and logistic regression was 0.59. CONCLUSION: Machine learning did not outperform logistic regression modeling in predicting postoperative complications after pancreatoduodenectomy.

The prediction of surgical complications using artificial intelligence in patients undergoing major abdominal surgery: A systematic review.

BACKGROUND: Conventional statistics are based on a simple cause-and-effect principle. Postoperative complications, however, have a multifactorial and interrelated etiology. The application of artificial intelligence might be more accurate to predict postoperative outcomes. The objective of this study was to determine the current quality of studies describing the use of artificial intelligence in predicting complications in patients undergoing major abdominal surgery. METHODS: A literature search was performed in PubMed, Embase, and Web of Science. Inclusion criteria were (1) empirical studies including patients undergoing (2) any type of gastrointestinal surgery, including hepatopancreaticobiliary surgery, whose (3) complications or mortality were predicted with the use of (4) any artificial intelligence system. Studies were screened for description of method of validation and testing in methodology. Outcome measurements were sensitivity, specificity, accuracy, and area under the receiver operating characteristic curve. RESULTS: From a total of 1,537 identified articles, 15 were included for the review. Among a large variety of algorithms used by the included studies, sensitivity was between 0.06 and 0.96, specificity was between 0.61 and 0.98, accuracy was between 0.78 and 0.95, and area under the receiver operating characteristic curve varied between 0.50 and 0.96. CONCLUSION: Artificial intelligence algorithms have the ability to accurately predict postoperative complications. Nevertheless, algorithms should be properly tested and validated, both internally and externally. Furthermore, a complete database and the absence of unsampled imbalanced data are absolute prerequisites for algorithms to predict accurately.

The muscle fiber type-fiber size paradox: hypertrophy or oxidative metabolism?

An inverse relationship exists between striated muscle fiber size and its oxidative capacity. This relationship implies that muscle fibers, which are triggered to simultaneously increase their mass/strength (hypertrophy) and fatigue resistance (oxidative capacity), increase these properties (strength or fatigue resistance) to a lesser extent compared to fibers increasing either of these alone. Muscle fiber size and oxidative capacity are determined by the balance between myofibrillar protein synthesis, mitochondrial biosynthesis and degradation. New experimental data and an inventory of critical stimuli and state of activation of the signaling pathways involved in regulating contractile and metabolic protein turnover reveal: (1) higher capacity for protein synthesis in high compared to low oxidative fibers; (2) competition between signaling pathways for synthesis of myofibrillar proteins and proteins associated with oxidative metabolism; i.e., increased mitochondrial biogenesis via AMP-activated protein kinase attenuates the rate of protein synthesis; (3) relatively higher expression levels of E3-ligases and proteasome-mediated protein degradation in high oxidative fibers. These observations could explain the fiber type-fiber size paradox that despite the high capacity for protein synthesis in high oxidative fibers, these fibers remain relatively small. However, it remains challenging to understand the mechanisms by which contractile activity, mechanical loading, cellular energy status and cellular oxygen tension affect regulation of fiber size. Therefore, one needs to know the relative contribution of the signaling pathways to protein turnover in high and low oxidative fibers. The outcome and ideas presented are relevant to optimizing treatment and training in the fields of sports, cardiology, oncology, pulmonology and rehabilitation medicine.

The Predictive Value of the Load Sharing Classification Concerning Sagittal Collapse and Posterior Instrumentation Failure: A Systematic Literature Review.

STUDY DESIGN: Systematic review. OBJECTIVE: In 1994, the Load Sharing Classification (LSC) was introduced to aid the choice of surgical treatment of thoracolumbar spine fractures. Since that time this classification system has been commonly used in the field of spine surgery. However, current literature varies regarding its use and predictive value in relation to implant failure and sagittal collapse. The objective of this study is to assess the predictive value of the LSC concerning the need for anterior stabilization to prevent sagittal collapse and posterior instrumentation failure. METHODS: An electronic search of PubMed, Medline, Embase, and the Cochrane Library was performed. Inclusion criteria were (1) cohort or clinical trial (2) including patients with thoracolumbar burst fractures (3) whose severity of the fractured vertebrae was assessed by the LSC. RESULTS: Five thousand eighty-two articles have been identified, of which 21 articles were included for this review. Twelve studies reported no correlation between the LSC and sagittal collapse or instrumentation failure in patients treated with short-segment posterior instrumentation (SSPI). Seven articles found no significant relation; 5 articles found no instrumentation failure at all. The remaining 9 articles experienced failure in patients with a high LSC or recommended a different surgical technique. CONCLUSIONS: Although the LSC was originally developed to predict the need for anterior stabilization in addition to SSPI, many studies show that SSPI only can be sufficient in treating thoracolumbar fractures regardless of the LSC. The LSC might have lost its value in predicting sagittal collapse and posterior instrumentation failure.

Implementation of robot-assisted Ivor Lewis procedure: Robotic hand-sewn, linear or circular technique?

BACKGROUND: Robot-assisted surgery for esophageal cancer is increasingly applied. Despite this upsurge, the preferential technique to create a robot-assisted intrathoracic anastomosis has not been established. DATA SOURCES: Bibliographic databases were searched to identify studies that performed a robot-assisted Ivor Lewis esophagectomy and described the technical details of the anastomotic technique. Out of 1701 articles, 16 studies were included for systematic review. CONCLUSIONS: This review shows that all technique used to create a thoracoscopic anastomosis can be adopted to robotic surgery. Techniques can be divided into three categories: robotic hand-sewn, circular stapling or linear stapling and robotic hand-sewn closure of the stapler defect. With limited robotic experience, circular stapling might be the preferred technique, however requires a well-trained bedside assistant. The linear stapling technique or hand-sewn technique are more challenging but enable experienced robotic surgeons to perform a controlled anastomosis without bedside support.

Postnatal development of fiber type composition in rabbit jaw and leg muscles.

We examined the difference in fiber type composition and cross-sectional areas during postnatal development in male rabbit jaw muscles and compared these with changes in leg muscles. The myosin heavy chain (MyHC) content of the fibers was determined by immunohistochemistry. No fiber type difference was found between the jaw muscles in 20-week-old rabbits. However, the way this adult fiber type composition was reached differed between the muscles. The deep temporalis, medial pterygoid, and superficial masseter displayed an increase in alpha fibers during early and a decrease during late postnatal development. Other jaw muscles displayed an increase in alpha fibers during early development only. In contrast, alpha fibers were not found in the soleus, in which fiber type changes were completed at week 4. The gastrocnemius muscle did not change its fiber type composition. Initially, fibers in jaw-opening muscles had larger cross-sectional areas than in other muscles, but they increased less during development. Although there were no large differences in the fiber type composition of muscles in young adult rabbits, large differences were found in the jaw muscles, but not in the leg muscles, during development. In part, these developmental changes in fiber percentages within the jaw muscles can be explained by functional modifications in this muscle group. In the present study, the deep temporalis, medial pterygoid, and superficial masseter showed the most dramatic percent changes in fibers during postnatal development.

A validation analysis of two self-reported HAM-D6 versions.

OBJECTIVE: The six items of the clinician-administrated Hamilton Depression Scale (HAM-D(6)) cover the core items of depressive states reflecting the antidepressive effect of medication. In this study, the two self-reported versions of the HAM-D(6) have been psychometrically validated to ensure the unidimensionality of this administration form in patients with mild-to-moderate depression. METHOD: The item response theory analysis of Mokken was used to test the unidimensionality of both the Interactive Voice Recording System (IVRS) version of the HAM-D(6) and a paper-and-pencil self-reported version (S-HAM-D(6)). Patients with typical major depression and with seasonal affective disorder were included. RESULTS: The Mokken analysis showed that the two self-reported versions of the HAM-D(6) obtained coefficients of homogeneity above 0.40, similar to the clinician-rated HAM-D(6) and thus implying unidimensionality. By contrast, the full HAM-D(17) versions (self-reported as well as clinician-rated) obtained coefficients of homogeneity below 0.40, implying that the HAM-D(17) is a multidimensional scale. CONCLUSION: The analysis show that both the IVRS version and the S-HAM-D(6) version are unidimensional self-rating scales for the measurement of depressive states.

Is fiber-type composition related to daily jaw muscle activity during postnatal development?

AIM: Muscles containing large numbers of slow-contracting fibers are generally more active than muscles largely composed of fast fibers. This relationship between muscle activity and phenotype suggests that (1) changes in fiber-type composition during postnatal development are accompanied by changes in daily activity and (2) individual variations in fiber-type composition are related to similar variations in daily muscle activity. METHODS: The masseter and digastric muscles of 23 New Zealand White rabbits (young, juvenile and adult) were examined for their phenotype (myosin heavy chain content) and their daily activity (total daily number of activity bursts). RESULTS: During development, the masseter showed a strong increase in the number of fast-type fibers compared to the number of slow-type fibers. During development, also the number of powerful bursts in the masseter increased. The digastric showed no significant changes in fiber types or burst numbers. Within each muscle, across individual animals, no significant correlations (R < 0.70) were found between any of the fiber types and daily burst numbers in any of the age groups. CONCLUSIONS: The results suggest that activity-related influences are of relatively minor importance during development and that other factors are dominant in determining fiber-type composition.

Daily activity of the rabbit jaw muscles during early postnatal development.

Early postnatal development of the jaw muscles is characterized by the transition from suckling to chewing behavior. As chewing develops the jaw closing muscles become more powerful compared with the jaw openers. These changes are likely to affect the amount of daily muscle activity. Therefore, the purpose of this study was to characterize for a jaw opener (digastric) and jaw closer (masseter) the total duration of daily muscle activity (i.e. the duty time), and the daily burst numbers and lengths during early postnatal development. Using radiotelemetry the activity of these muscles was recorded in 10 young New Zealand White rabbits between three and eight weeks of age. Fiber-type composition was analyzed at eight weeks of age by determining the myosin heavy chain content of the fibers. During postnatal development both muscles showed no significant decrease or increase in their daily activity. However, the interindividual variation of the duty time and burst number significantly decreased. There were no significant differences between the digastric and masseter except for the most powerful activities at eight weeks of age, where the masseter showed a significantly higher duty time and burst number than the digastric. The masseter contained a higher number of slow-type fibers expressing myosin heavy chain-I and myosin heavy chain-cardiac alpha than the digastric. The present results suggest that the amount of jaw muscle activation is already established early during postnatal development, before the transition from suckling to chewing behavior. This amount of activation seems to be related to the number of slow-type fibers.

Lubeluzole in acute ischemic stroke treatment: A double-blind study with an 8-hour inclusion window comparing a 10-mg daily dose of lubeluzole with placebo.

BACKGROUND AND PURPOSE: This trial was a double-blind, placebo-controlled, phase III trial with an 8-hour inclusion window to assess the efficacy and safety of an intravenous loading dose of 7.5 mg followed by a daily intravenous dose of 10 mg lubeluzole for 5 days in acute ischemic stroke patients. METHODS: A total of 1786 patients were randomized: 901 to lubeluzole and 885 to placebo. Overall, 212 patients (23.5%) from the lubeluzole group and 213 (24.1%) from the placebo group discontinued the trial prematurely. In the lubeluzole group 201 patients (22.3%) discontinued because of adverse events compared with 193 patients (21.8%) in the placebo group. RESULTS: The primary population for the efficacy analysis comprised the core stroke patients (exclusion of older patients aged >75 years with severe stroke) in the 0- to 6-hour inclusion time window. The primary efficacy parameter was a 3-category functional status (Barthel Index 70 to 100/0 to 70/vegetative, dead) at week 12. In the lubeluzole group 207 patients (47.8%) were classified as mildly dependent/independent at week 12, 131 (30.3%) were moderately/severely dependent, and 95 (21.9%) were vegetative/dead. In the placebo group these numbers were 221 (54.4%), 112 (27.6%), and 73 (18.0%), respectively. Logistic regression analysis showed no statistically significant difference between the treatment groups (P:=0.162). Additionally, for none of the secondary efficacy parameters (mortality at week 12, modified Rankin score, total Barthel score) was a statistically significant difference between the lubeluzole and placebo groups obtained. There were no statistically significant differences between the 2 treatments for all treated patients, patients included within the 6- to 8-hour window, and patients with severe strokes aged >75 years. Overall, of all treated patients, 401 (22.5%) died: 203 (22.5%) in the lubeluzole group and 198 (22.4%) with placebo. Of all subjects treated, 853 (95%) on lubeluzole and 826 (93%) on placebo reported an adverse event during their treatment period or within the next 2 days after discontinuation of treatment. The most frequently observed adverse events were fever (25.9% lubeluzole; 23.4% placebo), constipation (20.2%; 19.7%), and headache (17.6%; 21.2%). Imbalances were found for atrial fibrillation (1.8% lubeluzole; 1.1% placebo) and QT prolongation (0.9%; 0.2%). CONCLUSIONS: This study failed to show an efficacy of lubeluzole in the treatment of acute stroke. On the other hand, lubeluzole treatment by the current dosage schedule was not associated with a significant safety problem.

Galantamine in AD: A 6-month randomized, placebo-controlled trial with a 6-month extension. The Galantamine USA-1 Study Group.

BACKGROUND: Galantamine is a reversible, competitive cholinesterase inhibitor that also allosterically modulates nicotinic acetylcholine receptors. These mechanisms of action provided the rationale for a therapeutic trial of galantamine in AD. METHODS: A 6-month, multicenter, double-blind trial was undertaken in 636 patients with mild to moderate AD. Patients were randomly assigned to placebo or galantamine and escalated to maintenance doses of 24 or 32 mg/d. Eligible patients then entered a 6-month, open-label study of the 24 mg/d dose. Primary efficacy measures were the 11-item AD Assessment Scale cognitive subscale (ADAS-cog/11) and the Clinician's Interview-Based Impression of Change plus Caregiver Input (CIBIC-plus). The Disability Assessment for Dementia (DAD) scale was a secondary efficacy variable. RESULTS: Galantamine significantly improved cognitive function relative to placebo; the treatment effects were 3.9 points (lower dose) and 3.8 points (higher dose) on the ADAS-cog/11 scale at month 6 (p < 0.001 in both cases). Both doses of galantamine produced a better outcome on CIBIC-plus than placebo (p < 0.05). Therapeutic response to galantamine was not affected by APOE genotype. At 12 months, mean ADAS-cog/11 and DAD scores had not significantly changed from baseline for patients who received galantamine 24 mg/d throughout the 12 months. The most common adverse events, which were predominantly gastrointestinal, decreased in frequency during long-term treatment. There was no evidence of hepatotoxicity. CONCLUSIONS: Galantamine is effective and safe in AD. At 6 months, galantamine significantly improved cognition and global function. Moreover, cognitive and daily function were maintained for 12 months with the 24 mg/d dose.

Expression of catecholamine-synthesizing enzymes, peptidylglycine alpha-amidating monooxygenase, and neuropeptide Y mRNA in the rat adrenal medulla after acute systemic nicotine.

The expression of catecholamine-synthesizing enzymes in the adrenal medulla is upregulated in parallel by stress and pharmacological treatments. In this study we examined whether a neuropeptide and its processing enzyme are regulated in parallel with catecholamine enzyme genes after drug treatment. Because the main effect of stress on the adrenal medulla is via splanchnic nerve stimulation of nicotinic receptors, we used nicotine to stimulate the medulla and visualized expression of catecholamine enzyme genes, the medullary peptide neuropeptide Y (NPY), and the neuropeptide-processing enzyme peptidylglycine alpha-amidating monooxygenase (PAM) by in situ hybridization quantified by image analysis of autoradiographic images. Rats received a single injection of nicotine (0, 1, or 5 mg/kg sc). Six hours later, rats were transcardially perfused. Free-floating adrenal gland sections were hybridized with 35S-labeled cDNA probes for tyrosine hydroxylase (TH), dopamine beta-hydroxylase (DBH), phenylethanolamine N-methyltransferase (PNMT), PAM, and NPY. Nicotine treatment upregulated the expression of TH, PNMT, and NPY genes in a dose-dependent fashion. Small but nonsignificant increases were observed in DBH and PAM mRNA levels. These results suggest that common transcriptional activation mechanisms may upregulate both catecholamine and neuropeptide synthesis in the adrenal medulla after nicotinic stimulation.

Parallel upregulation of catecholamine-synthesizing enzymes in rat brain and adrenal gland: effects of reserpine and correlation with immediate early gene expression.

Changes in the mRNA levels of all catecholamine-synthesizing enzymes were examined 24 h after a single injection of reserpine by in situ hybridization. The responses of the midbrain dopaminergic cells in the ventral tegmental area and substantia nigra compacta, locus ceruleus and adrenal gland were studied in three groups of animals receiving either no injection, vehicle injection or reserpine 10 mg/kg subcutaneously. Increases in enzyme message signal observed by in situ hybridization were corroborated by Northern blot analysis for all four enzyme mRNAs species expressed in the locus ceruleus and adrenal gland were found while no change of enzyme message was detected the midbrain. Two distinct subpopulations of adrenomedullary cells could be distinguished by their baseline levels of enzyme mRNA expression: the majority of medullary cells have moderate adrenomedullary cells could be distinguished by their baseline levels of enzyme mRNA expression: the majority of medullary cells have moderate levels of all four enzyme mRNAs but a minority of cells show very high signal for the first three enzymes of the catecholamine synthesis pathway. To test whether reserpine elicits a selective transcriptional response of the catecholamine enzyme genes or induces other neuronal genes, cDNA probes for the growth-associated protein GAP-43 which is highly expressed and neurofilament L which is weakly expressed in monoaminergic neurons were used as independent cellular markers and showed no change in message levels. Changes in mRNA levels of the proto-oncogenes c-fos and c-jun were examined 1 h after injection of reserpine by in situ hybridization and compared to the pattern observed for the Fos protein immunohistochemically. C-fos and c-jun proto-oncogene activation was observed 1 h after reserpine in the locus ceruleus and adrenal medulla, specifically in those catecholaminergic structures that respond with increased enzyme gene transcription; in contrast, the dopaminergic neurons of the substantia nigra did not exhibit detectable proto-oncogene activation, only a small group of neurons in the ventral tegmental area showed c-fos without concomitant c-jun expression after reserpine.

Axotomy-induced differential gene induction in neurons of the locus ceruleus and substantia nigra.

The biochemical and molecular events correlated with neuronal injury and survival are not well understood. Previous studies have reported that following axotomy, neurons in the substantia nigra pars compacta (SNc) and the locus ceruleus (LC) exhibit a significant increase in tyrosine hydroxylase (TH) enzyme activity within 24-72 h (Brain Res., 144 (1978) 325-342; Brain Res., 92 (1975) 57-72). To investigate the potential contribution of TH gene induction to this increase a semi-quantitative immunocytochemical and in situ hybridization time course analysis was undertaken. Following axotomy, TH immunoreactivity increased in neurons of both the SNc and LC. In contrast, an increase in TH mRNA was only evident in neurons of the LC. As a possible mechanism for the observed alterations in TH gene expression, the levels of an immediate early gene, c-fos, were examined. C-fos mRNA and Fos protein were not expressed in either normal or axotomized neurons of the SNc. However, the constitutive expression in control LC neurons increased significantly following axotomy. These data demonstrate the differential response of two central catecholaminergic populations to axotomy and suggests a potential role for the immediate early gene, c-fos, in the post-injury reaction.

Decrease in tyrosine hydroxylase, but not aromatic L-amino acid decarboxylase, messenger RNA in rat olfactory bulb following neonatal, unilateral odor deprivation.

Unilateral naris cauterization in rats results in occlusion of the affected naris and blockade of odorant access to ipsilateral olfactory receptor cells in the olfactory epithelium. These receptor cells project exclusively to the olfactory bulb (OB) and appear to regulate expression of the dopaminergic phenotype in a population of OB juxtaglomerular neurons. Unilateral odor deprivation results in a loss of normal stimulatory input to the OB and a marked and specific decrease in ipsilateral OB tyrosine hydroxylase (TH) expression. The expression of co-localized aromatic L-amino acid decarboxylase (AADC) is not similarly affected. We have used this procedure in neonatal rats to examine the effect of stimulus deprivation on OB TH and AADC mRNA levels. Both Northern blot and in situ hybridization analyses revealed a pronounced decrease in ipsilateral as compared to contralateral OB TH mRNA levels 40 days after naris closure. In contrast, the levels of OB AADC mRNA were unaltered by naris closure. By in situ hybridization histochemistry, both TH and AADC mRNAs were localized to OB juxtaglomerular neurons. Odor deprivation was associated with an apparent region-specific reduction in TH mRNA within the ipsilateral OB glomerular layer. By densitometric analysis, the loss of TH-specific message was quantitatively consistent with the decrease in TH activity, suggesting that the observed plasticity of OB dopaminergic neurons following functional deafferentation can be attributed to a selective, transneuronally-mediated down regulation of TH gene transcription.

Characterization of recombinant bovine phenylethanolamine N-methyltransferase expressed in a mouse C127 cell line.

Bovine phenylethanolamine N-methyltransferase (PNMT) cDNA was inserted into a bovine papilloma virus-based expression vector and used to transfect a mouse C127 cell line. The resultant recombinant bovine PNMT was characterized biochemically and immunochemically. Recombinant bovine PNMT activity, like the native bovine enzyme, was enhanced by phosphate ion in a concentration-dependent manner. Their molecular weights were shown to be identical by Western blot analysis. Antibodies raised against native bovine adrenal PNMT equally immunoprecipitated the activity of the recombinant and native enzymes. In addition, double immunodiffusion analysis showed a single precipitin line of confluence with both enzyme preparations, indicating immunological identity of native and recombinant bovine PNMT. These antibodies immunostained the recombinant enzyme protein in transfected cells and in their neurite-like processes. In addition, in situ hybridization with the bovine PNMT cDNA probe resulted in a labelling pattern similar to the immunostaining. The recombinant bovine PNMT as the native bovine enzyme exist in multiple-charge forms, but only one form is predominant. Taken together, our results suggest that recombinant bovine PNMT, expressed from bovine PNMT cDNA in a mouse cell line is enzymatically active and shares many common features with native bovine adrenal PNMT.