Pancreatic exocrine insufficiency following pancreatoduodenectomy: A prospective bi-center study.

BACKGROUND/OBJECTIVES: Pancreatic exocrine insufficiency (PEI) is a common complication following pancreatoduodenectomy (PD) leading to malnutrition. The course of PEI and related symptoms and vitamin deficiencies is unknown. This study aimed to assess the (long-term) incidence of PEI and vitamin deficiencies after PD. METHODS: A bi-centre prospective observational cohort study was performed, including patients who underwent PD for mainly pancreatic and periampullary (pre)malignancies (2014-2018). Two cohorts were formed to evaluate short and long-term results. Patients were followed for 18 months and clinical symptoms were evaluated by questionnaire. PEI was based on faecal elastase-1 (FE-1) levels and/or clinical symptoms. RESULTS: In total, 95 patients were included. After three months, all but three patients had developed PEI and 27/29 (93%) patients of whom stool samples were available showed abnormal FE-1 levels, which did not improve during follow-up. After six months, all patients had developed PEI. During follow-up, symptoms resolved in 35%-70% of patients. Vitamin D and K deficiencies were observed in 48%-79% of patients, depending on the moment of follow-up; 0%-50% of the patients with deficiencies received vitamin supplementation. DISCUSSION: This prospective study found a high incidence of PEI after PD with persisting symptoms in one-to two thirds of all patients. Limited attention was paid to vitamin deficiencies. Improved screening and treatment strategies for PEI and vitamins need to be designed.

Biosecurity Assessment and Seroprevalence of Respiratory Diseases in Backyard Poultry Flocks Located Close to and Far from Commercial Premises.

Raising backyard chickens is an ever-growing hobby in the United States. These flocks can be a substrate for respiratory disease amplification and transmission to commercial facilities. Five hundred fifty-four chickens from 41 backyard flocks were sampled in this study. ELISA kits were used to detect antibodies against avian influenza (AI), infectious laryngotracheitis (ILT), Newcastle disease (ND), infectious bronchitis (IB), Ornithobacterium rhinotracheale (ORT), Mycoplasma gallisepticum (MG), and Mycoplasma synoviae (MS). All visited flock owners answered a biosecurity questionnaire that assessed biosecurity measures. The questionnaire revealed that backyard poultry owners lack simple biosecurity measures such as use of dedicated shoes, their chicken sources are unreliable, and few of them benefit from veterinary oversight. Only one flock had a clear vaccination history against ND and IB. ORT, ND, IB, MS, MG, and ILT were the most seroprevalent in backyard poultry flocks with 97% (41/42), 77.5% (31/40), 75% (30/40), 73% (31/42), 69% (29/42), and 45% (19/42), respectively. The vaccinated flock was not considered in these calculations. When examining the distance between backyard flocks and the nearest commercial poultry facility, ND and MG were significantly more likely to be found in backyard flocks close to (<4 miles) whereas ORT was significantly more likely in backyard chickens located far from (>4 miles) commercial poultry. Birds purchased directly from National Poultry Improvement Plan hatcheries showed a reduced ND, MG, and MS antibody prevalence. Wearing dedicated shoes decreased MS antibody-positive birds. Finally, history of wild bird contact had a clear effect on an increased seroprevalence of NDV and MG. Serological results suggest that backyard poultry flocks have the potential to serve as a reservoir or amplifier for poultry respiratory diseases. The information generated in this project should direct extension efforts toward emphasizing the importance of small flock biosecurity and chick acquisition sources.

The effect of side of implantation on unilateral cochlear implant performance in patients with prelingual and postlingual sensorineural hearing loss: A systematic review.

OBJECTIVE: Cerebral lateralisation of language processing leads to a right ear advantage in normal hearing subjects. The aim of this study was to present a systematic overview of the effect of implantation side on postoperative cochlear implant performance in patients with symmetrical severe to profound sensorineural hearing loss. DATA SOURCES: PubMed, Embase and The Cochrane Library databases. RESEARCH METHODS: Databases were searched from database inception up to 9 January 2017 for cochlear implant and side and all synonyms. Title, abstract and full-text of retrieved articles were screened for eligibility. Then, directness of evidence and risk of bias were assessed. For the included articles, study characteristics and outcome data (hearing and language development) were extracted. RESULTS: 2541 unique articles were screened, of which twenty were eligible for critical appraisal. No randomised controlled trials were identified. Twelve studies with a high directness of evidence remained for data extraction. Four of six studies including children with pre-lingual sensorineural hearing loss and four of seven studies investigating adults with postlingual sensorineural hearing loss found a right ear advantage in at least one outcome measurement related to cochlear implant performance. CONCLUSION: The available evidence on the effect of side of implantation is of low quality, as study populations and outcome measures are heterogeneous. The majority of studies reveals evidence for a right ear advantage in prelingually deafened children as well as postlingually deafened adults. In view of the present evidence and as no left ear advantage was identified, we cautiously advise implanting the cochlear implant in the right ear when other prognostic factors do not favour the left ear and sensorineural hearing loss is symmetrical.

In vivo treatment of hemophilia A and mucopolysaccharidosis type VII using nonprimate lentiviral vectors.

Gene therapy holds great promise for the treatment of a variety of inherited diseases, including hemophilia A and mucopolysaccharidosis type VII (MPS VII). In both these disorders, subnormal levels of replacement protein have therapeutic effects. Thus we hypothesized that transduction of a small proportion of cells by feline immunodeficiency virus (FIV)-based lentiviral vectors might provide sufficient levels of transgene expression for phenotypic correction. We intravenously injected replication-deficient FIV-based vectors encoding either human factor VIII or human beta-glucuronidase into factor VIII-deficient or beta-glucuronidase-deficient mice, respectively. This route of delivery targeted multiple organs, with the liver as the primary transduction site. In the hemophilia A mice, factor VIII expression persisted for the duration of the experiments (approximately 5 months), and recipient mice survived an otherwise lethal bleeding episode (tail-clipping). In mucopolysaccharidosis type VII mice, substantial beta-glucuronidase activity was detected in several tissues and corresponded with marked reduction of lysosomal storage in liver and spleen. These findings indicate that gene transfer with FIV-based lentiviral vectors can permanently introduce transgenes into a sufficient number of hepatocytes for long-term therapeutic effect and suggest potential clinical value of FIV-based lentiviral vectors for treatment of hemophilia A and MPS VII.

Recombinant adeno-associated virus type 2, 4, and 5 vectors: transduction of variant cell types and regions in the mammalian central nervous system.

Recombinant adeno-associated virus vectors based on serotype 2 (rAAV2) can direct transgene expression in the central nervous system (CNS), but it is not known how other rAAV serotypes perform as CNS gene transfer vectors. Serotypes 4 and 5 are distinct from rAAV2 and from each other in their capsid regions, suggesting that they may direct binding and entry into different cell types. In this study, we examined the tropisms and transduction efficiencies of beta-galactosidase-encoding vectors made from rAAV4 and rAAV5 compared with similarly designed rAAV2-based vectors. Injection of rAAV5 beta-galactosidase (betagal) or rAAV4betagal into the lateral ventricle resulted in stable transduction of ependymal cells, with approximately 10-fold more positive cells than in mice injected with rAAV2betagal. Major differences between the three vectors were revealed upon striatal injections. Intrastriatal injection of rAAV4betagal resulted again in striking ependyma-specific expression of transgene, with a notable absence of transduced cells in the parenchyma. rAAV2betagal and rAAV5betagal intrastriatal injections led to beta-gal-positive parenchymal cells, but, unlike rAAV2betagal, rAAV5betagal transduced both neurons and astrocytes. The number of transgene-positive cells in rAAV5betagal-injected brains was 130 and 5,000 times higher than in rAAV2betagal-injected brains at 3 and 15 wk, respectively. Moreover, transgene-positive cells were widely dispersed throughout the injected hemisphere in rAAV5betagal-transduced animals. Together, our data provide in vivo support for earlier in vitro work, suggesting that rAAV4 and rAAV5 gain cell entry by means of receptors distinct from rAAV2. These differences could be exploited to improve gene therapy for CNS disorders.

Is ethanol an important antioxidant in alcoholic beverages associated with risk reduction of cataract and atherosclerosis?

It has been reported in the epidemiological literature that cataract, stroke, and atherosclerosis risk is reduced by 50% in people consuming one alcoholic drink per day. Peroxide has been implicated as a causative agent in cataractogenesis, and LDL oxidation appears to play a role in atherosclerosis. The antioxidant activity of alcohol was measured by: (i) use of a luminescent assay developed in our laboratory, confirmed as appropriate; (ii) electron spin resonance (ESR) spin-trapping; and (iii) copper-catalysed oxidation of LDL and VLDL from hamsters fed 6% ethanol in their drinking water. Ethanol reduced the luminescent counts/min from peroxide and superoxide. It significantly reduced the spin-trapped signal of hydroxyl radical, but not the superoxide signal. Other alcohols also showed large reductions in counts from hydrogen peroxide. Plasma from hamsters fed 6% ethanol had lower lipid peroxides and the oxidizability of LDL and VLDL was significantly reduced compared to controls. These data provide a possible explanation for the effect of beverages containing ethanol in the reduction of cataract and atherosclerosis risk observed in human population studies.

Intracellular trafficking of the JNCL protein CLN3.

Juvenile neuronal ceroid lipofuscinosis is a lysosomal storage disease that causes visual impairment, progressive mental deterioration, and eventually death. A predominant 1.02-kb deletion as well as other mutations have been described in the CLN3 gene. Lacking significant identity with proteins of known function and no overt targeting signals within the primary amino acid sequence, accurate predictions of the intracellular location and function could not be made. Further, recent conflicting reports identified CLN3 as either a lysosomal or a mitochondrial protein. Transfection experiments using native and epitope-tagged fusion proteins were evaluated to help delineate CLN3 localization. We confirmed by immunohistochemistry and brefeldin A treatment that NH2-terminal green fluorescence protein (GFP)-CLN3 fusion proteins were retained in the Golgi apparatus, with no colocalization with mitochondrial markers. Anti-CLN3 antibodies directed against amino acids 67-90 of CLN3 were generated and shown to be specific for a 50-kDa protein in HEK 293 cells and GFP-CLN3 in transfected cells. However, cells transfected with nontagged CLN3 or carboxyl-terminal-tagged CLN3 were not immunoreactive with anti-CLN3 antibodies, suggesting that normally, the amino terminus interacts with other molecules. Thus, tags on the NH2-terminus probably inhibited these interactions and movement of CLN3 from the Golgi to more distal compartments. Also, CLN3 tagged at the COOH-terminus with either GFP or FLAG epitopes were retained in the ER, indicating a role for the COOH-terminus in trafficking. Taken together, these data confirm that CLN3 traffics through the ER and Golgi.

Systemic and central nervous system correction of lysosomal storage in mucopolysaccharidosis type VII mice.

Mucopolysaccharidosis (MPS) type VII patients lack functional beta-glucuronidase, leading to systemic and central nervous system dysfunction. In this study we tested whether recombinant adenovirus that encodes beta-glucuronidase (Adbetagluc), delivered intravenously and into the brain parenchyma of MPS type VII mice, could provide long-term transgene expression and correction of lysosomal distension. We also tested whether systemic treatment with the immunosuppressive anti-CD40 ligand antibody, MR-1, affected transgene expression. We found substantial plasma beta-glucuronidase activity for over 9 weeks after gene transfer in the MR-1- treated group, with subsequent decline in activity corresponding to a delayed anti-beta-glucuronidase antibody response. At 16 weeks, near wild-type amounts of beta-glucuronidase activity and striking reduction of lysosomal pathology were detected in livers from mice that had received either MR-1 cotreatment or control antibody. In the lung and kidney, beta-glucuronidase activity was markedly higher for the MR-1-treated group. beta-Glucuronidase activity in the brain persisted independently of MR-1 treatment. Activity was intense in the injected hemisphere and was also evident in the noninjected cortex and striatum, with dramatic improvements in storage deposits in areas of both hemispheres. These results indicate that prolonged enzyme expression from transgenes delivered to deficient liver and brain can mediate pervasive correction and illustrate the potential for gene therapy of MPS and other lysosomal storage diseases.

Systemic hyperosmolality improves beta-glucuronidase distribution and pathology in murine MPS VII brain following intraventricular gene transfer.

Mucopolysaccharidosis VII, a classical lysosomal storage disease, is caused by deficiency of the enzyme beta-glucuronidase. Central nervous system (CNS) manifestations are severe with accumulations of storage vacuoles in all cell types. Intraventricular gene transfer can lead to transduction of the ependyma, with production and secretion of beta-glucuronidase into the cerebral spinal fluid and underlying cortex resulting in reversal of disease pathology restricted to the periventricular areas. We tested if systemic hyperosmolality would increase the distribution of beta-glucuronidase in brain parenchyma after intraventricular virus injection. Mice were administered mannitol, intraperitoneally, 20 days after gene transfer and 1 day prior to sacrifice. Mannitol-induced systemic hyperosmolality caused a marked penetration of beta-glucuronidase into the brain parenchyma. If mannitol was administered at the time of the intraventricular injection of virus, there was penetration of vector across the ependymal cell layer, with infection of cells in the subependymal region. This also resulted in increased beta-glucuronidase activity throughout the brain. Sections of brains from beta-glucuronidase-deficient mice showed correction of cellular pathology in the subependymal region plus cortical structures away from the ventricular wall. These data indicate that virus-mediated gene transfer to the brain via the ventricles, coupled with systemic mannitol administration, can lead to extensive CNS distribution of beta-glucuronidase with concomitant correction of the storage defect. Our findings have positive therapeutic implications for the treatment of CNS disorders with gene transfer vectors and recombinant proteins.

Extensive beta-glucuronidase activity in murine central nervous system after adenovirus-mediated gene transfer to brain.

Mucopolysaccharidosis type VII (MPS VII), caused by beta-glucuronidase deficiency, is a classic lysosomal storage disease. In the central nervous system (CNS), there is widespread pathology with distention of vacuoles in neurons and glia. An approach to therapy for MPS VII would require extensive delivery of enzyme to the CNS and subsequent uptake by the affected cells. In this study we show that intrastriatal injection of recombinant adenovirus encoding beta-glucuronidase (Ad betagluc) to MPS VII or wild-type mice results in focal, intense beta-glucuronidase mRNA expression near the injection site. Further, histochemical staining for enzyme activity showed that beta-glucuronidase activity extended well beyond transduced cells. Activity was detected throughout the ipsilateral striatum as well as in the corpus callosum, ventricles, and bilateral neocortex. Similarly, after injection into the right lateral ventricle or cisterna magna, enzyme activity was present in the ependymal cells of the ventricles, in the subarachnoid spaces, and also in the underlying cortex (150-500 microm from ependyma). The distribution of enzyme was most extensive 21 days after gene transfer to normal mouse brain, with more than 50% of the hemisphere positive for beta-glucuronidase activity. Eighty-four days after adenovirus injection a substantial level of enzyme expression remained (>40% of hemisphere positive for beta-glucuronidase activity). Histological sections from striatum of beta-glucuronidase-deficient mice injected with Ad betagluc showed a marked reduction in the number of distended vacuoles in both neurons and glia, as compared with uninjected striatum. Importantly, correction was noted in both hemispheres. Our finding that a relatively small number of transduced cells produce enzyme that reaches a large proportion of the CNS has favorable implications in developing direct gene transfer therapies for lysosomal storage disorders.