Monoclonal antibody production to human and bovine 2′:3′-cyclic nucleotide 3′-phosphodiesterase (CNPase): high-specificity recognition in whole brain acetone powders and conservation of sequence between CNP1 and CNP2

Monoclonal antibodies against human and bovine 2′:3′-cyclic nucleotide 3′-phosphodiesterase (CNPase) were generated by fusing FOX-NY myeloma cells with spleen cells from RBF/Dn mice previously immunized with the purified brain antigens. The enzyme isolated from bovine brain was quite basic, with an isoelectric point of 9.71 and both the bovine and human enzymes consisted of a closely spaced doublet at approximately 44 and 46 kDa on SDS-PAGE. Six monoclonals were identified as strongly recognizing the enzyme on both ELISA plates and on immunoblots of whole brain protein. Four monoclonals very weakly cross-reacted with guinea pig myelin basic protein. In contrast with two previous reports, some of our monoclonal antibodies did immunostain 2 or 3 protein bands in peripheral nerve, two bands closely corresponding to those immunostained in central nervous system (CNS) myelin, the Wolfgram protein fraction and in acetone powders of whole brain. Each of the 6 monoclonals reacting strongly on immunoblots recognized the enzyme in from 2 to 5 of the species examined (human, bovine, rat, mouse and rabbit). In addition, all 6 monoclonals that immunostained the enzyme in whole brain, myelin and Wolfgram protein immunoblots recognized both CNP1 (44 kDa) and CNP2 (46 kDa). The two closely spaced protein bands observed on SDS-PAGE and previously stained on immunoblots of CNS CNPase using polyvalent rabbit anti-bovine CNPase antisera, and now different monoclonal antibodies, appear to be immunologically related and to contain highly conserved sequences.     

Nocturnal hypoglycaemia in Type 1 diabetic patients,assessed with continuous glucose monitoring: frequency,duration and associations

Aims We quantified the occurrence and duration of nocturnal hypoglycaemia in individuals with Type 1 diabetes treated with continuous subcutaneous insulin infusion (CSII) or multiple‐injection therapy (MIT) using a continuous subcutaneous glucose sensor. Methods A microdialysis sensor was worn at home by 24 patients on CSII (mean HbA_1c 7.8 ± 0.9%) and 33 patients on MIT (HbA_1c 8.7 ± 1.3%) for 48 h. Occurrence and duration of nocturnal hypoglycaemia were assessed and using multivariate regression analysis, the association between HbA_1c, diabetes duration, treatment type (CSII vs. MIT), fasting and bedtime blood glucose values, total daily insulin dose and mean nocturnal glucose concentrations, and hypoglycaemia occurrence and duration was investigated. Results Nocturnal hypoglycaemia ≤ 3.9 mmol/l occurred in 33.3% of both the CSII‐ (8/24) and MIT‐treated patients (11/33). Mean (± sd ; median, interquartile range) duration of hypoglycaemia ≤ 3.9 mmol/l was 78 (± 76; 57, 23–120) min per night for the CSII‐ and 98 (± 80; 81, 32–158) min per night for the MIT‐treated group. Multivariate regression analysis showed that bedtime glucose value had the strongest association with the occurrence (P = 0.026) and duration (P = 0.032) of nocturnal hypoglycaemia. Conclusions Microdialysis continuous glucose monitoring has enabled more precise quantification of nocturnal hypoglycaemia occurrence and duration in Type 1 diabetic patients. Occurrence and duration of nocturnal hypoglycaemia were mainly associated with bedtime glucose value.     

Morphological and proliferative responses of cultured Schwann cells following rapid phagocytosis of a myelin-enriched fraction

Summary Cultured Schwann cells were found to phagocytose exogenously applied myelin membranes within 1 h. However, the resulting proliferative response required an additional 9 h of incubation. Treatment with ammonium chloride, a lysosomal inhibitor, delayed the appearance of the proliferative response to the myelin membranes by 12 h. Processing of myelin within the Schwann cells was followed by the appearance of immunocytochemically detectable myelin basic protein which was first visible at 4 h. Similar to the proliferative response, the appearance of immunoreactive material was delayed by the addition of ammonium chloride. Schwann cells were observed initially to ingest myelin fragments at their distal-most tips after which time the myelin phagosomes collected in the perinuclear region and fused with lysosomes. Phagocytic Schwann cells had a notable increase in Golgi membranes and microfilaments and contained widely dilated, rough endoplasmic reticulum cisternae. In purified cell cultures, Schwann cells phagocytosed myelin slower than macrophages, but displayed phagocytic abilities much greater than fibroblasts. The ability of cultured Schwann cells to phagocytose myelin rapidly suggests that these cells may aid in the breakdown and removal of myelin during Wallerian degeneration. These data further confirm the mitogenic effect of myelin and its possible role during nerve regeneration.     

"Humanizing" childbirth: the discovery and implementation of bonding theory

Recent changes in methods of childbirth attendance represent one example of an emerging emphasis on the humanization of medical treatment. Although many have observed this trend toward humanization, the process of medical accommodation--including rationales provided for change, the nature and limitations of humanization, and the consequences of "humanized" medicine--is not well understood. This paper explores the process of medical accommodation by focusing on the ways in which the issue of parent-infant bonding has contributed to the humanization of obstetric care. The bonding issue has provided a rationale for change in a specialty area facing criticisms on several fronts. Acknowledging consumer demands for change in traditional styles of maternity care, medical professionals have responded by offering alternative programs based largely in the conclusions of research on the attachment process. It is demonstrated that these medically proposed alternatives suffer from many organizational constraints and that the emphasis on bonding fostered by these new programs has potentially negative consequences.     

Sentinel Node Biopsy Should Not be Routine in Older Patients with ER-Positive HER2-Negative Breast Cancer Who Are Willing and Able to Take Hormone Therapy

Annals of Surgical Oncology - The SSO Choosing Wisely campaign recommended selective sentinel lymph node biopsy (SLNB) in clinically node-negative women aged ≥ 70 years with ER+ breast...     

Comparison of cyclic nucleotide and energy metabolism of intact mouse retina in situ and in vitro.

Whole mouse retina in situ contains approximately 1 μm adenosine 3′,5′-monophosphate (cyclic AMP) and about fivefold higher levels of guanosine 3′,5′-monophosphate (cyclic GMP). Light-adapted retinas have only half as much of both cyclic nucleotides as dark-adapted retinas.Both cyclic AMP and cyclic GMP levels are modified substantially when retinas are removed from the eye and incubated in physiologic buffers. Cyclic GMP levels increase in light- and dark-adapted retinas, but a differential concentration is always maintained, and after a few minutes of incubation it is even greater than that in in situ retinas. In contrast, the difference between cyclic AMP levels obvious in in situ light- and dark-adapted retinas is obscured during the initial minutes of incubation by a transient rise of cyclic AMP in light-adapted retinas, but then becomes apparent again when cyclic AMP levels fall after about 15 min of incubation. Cyclic AMP and cyclic GMP levels decrease rapidly when isolated dark-adapted retinas, in vitro, are exposed to light and the decrease is a function of light intensity. In contrast, cyclic GMP but not cyclic AMP levels increase when light-adapted retinas, in vitro, are placed in darkness.Ischemia causes a marked reduction of ATP and P-creatine levels and elevates cyclic AMP levels in light- and dark-adapted retinas, in situ. Ischemia depresses cyclic GMP levels in dark-adapted retinas, in situ, but has no effect in light-adapted retinas. In incubated retinas, oxygen deprivation produced by NaCN also decreases energy reserves; however, its effect on cyclic AMP is qualitatively similar to that in vivo only after 15 min of incubation. NaCN has little or no effect on cyclic GMP levels in isolated retinas.These data indicate that cyclic nucleotide regulation in intact, incubated retina has many similarities to that of retina, in vivo, and suggest that with due precautions retina maintained, in vitro, is a valid and useful experimental model system for biochemical and biophysical investigations.