Suspension culture of separated follicles consisting of differentiated thyroid epithelial cells

We have prepared thyroid follicles in suspension culture to use as a model system in vitro for investigation of some properties of the thyroid gland. The follicles were free of endothelial cells, fibroblasts, and other nonepithelial cells. They were prepared by collagenase treatment of minced rat thyroid glands followed by differential filtration of the suspension through nylon meshes. Small clusters of principal thyroid epithelial cells were separated from large fragments and single cells. They were cultured in dilute suspension in Coon's modified F-12 medium in dishes coated with agarose to avoid having the cells attach to the dishes. By culture day 3, most of the clusters formed closed follicles containing a periodic acid-Schiff-positive colloid but without a basal lamina. Follicle walls contained an occasional C cell. The epithelium resembled that in the thyroid of a recently hypophysectomized rat, with normal polarity and organelle complement normal with respect to position and abundance, with basally located lysosomes, no pseudopods, and no colloid droplets. The cells were responsive to thyroid-stimulating hormone (thyrotropin) and to dibutyryl cyclic AMP. Thyroid-stimulating hormone at 10 munits/ml resulted in apical migration of lysosomes and formation of pseudopods and colloid droplets within 30 min; longer exposure resulted in depletion of luminal colloid. The results indicate that the suspended follicles resemble follicles in vivo with respect to morphology and responsiveness to thyroid-stimulating hormone in the absence of other cell types.     

Differential recognition of a tyrosine-dependent signal in the basolateral and endocytic pathways of thyroid epithelial cells

Trafficking of receptors is of crucial importance for the physiology of most exocrine and endocrine organs. It is not known yet if the same mechanisms are used for sorting in the exocytic and endocytic pathways in the different epithelial tissues. In this work, we have used a deletion mutant of the human neurotrophin receptor p75(hNTR) that is normally localized on the apical membrane when expressed in Madin-Darby canine kidney cells. This internal 57-amino acid deletion of the cytoplasmic tail leads to a relocation of the protein from the apical to the basolateral membrane and to rapid and efficient endocytosis. These events are mediated by a signal localized within 9 amino acids of the mutated cytoplasmic tail that is strictly dependent on a tyrosine residue (Tyr-308). We have analyzed the basolateral sorting efficiency and endocytic capacity of this signal in Fischer rat thyroid (FRT) cells, in which basolateral and endocytic determinants have not yet been identified. We found that this targeting signal can mediate efficient transport to the basolateral membrane also in FRT cells with similar tyrosine dependence as in MDCK cells. In contrast to MDCK cells, this Tyr-based signal was not able to mediate coated pits localization and endocytosis in FRT cells. These data represent the first characterization of basolateral/endocytic signals in thyroid epithelial cells. Furthermore, our results indicate that requirements for tyrosine-dependent basolateral sorting signals are conserved among cell lines from different tissues but that the recognition of the colinear endocytic signal is tissue specific.     

Low serum bioactive luteinizing hormone in nonorganic male impotence: possible relationship with altered gonadotropin-releasing hormone pulsatility

We determined the biological activity of serum LH in 23 men, aged 25-50 yr, complaining of nonorganic impotence of at least 1-yr duration and 20 normal men. All of the impotent men had normal general physical examinations, penile Doppler tests, psychological tests, and peripheral nerve conduction. Serum PRL, FSH, LH, and thyroid hormone concentrations were normal as were the results of provocative tests of TSH, gonadotropin, and PRL secretion. The mean serum immunoreactive LH (I-LH) levels, measured in each impotent and normal man in three samples taken at 15-min intervals, were similar [7.2 +/- 0.5 (+/-SE) vs. 6.4 +/- 0.5 mIU/mL (IU/L)]. In contrast, the mean serum bioactive LH (B-LH) level was significantly lower in the impotent men than in the normal men [15.9 +/- 2.1 (+/-SE) vs. 33.0 +/- 2.8 mIU/mL (IU/L); P less than 0.05], as was the LH bio- to immunoactive (B/I) ratio (2.1 +/- 0.2 vs. 5.6 +/- 0.5; P less than 0.02). The mean serum testosterone level in the impotent men, although all individual values were within the range of normal for our laboratory [200-900 ng/100 mL (693-3120 nmol/L)], was 25% lower than that in the normal men [347 +/- 23 vs. 450 +/- 26 ng/100 mL; P less than 0.05 (1204 +/- 81 vs. 1560 +/- 91 nmol/L)]. In addition, a significant positive correlation was found between serum testosterone levels and LH B/I ratios in the impotent men (r = 0.45; P = 0.029). Pulsatile LH secretion, measured in six impotent and four normal men in blood samples collected every 15 min for 6 h, was similar in the two groups. The mean serum I-LH levels were similar [7.5 +/- 1.1 (+/-SE) vs. 5.1 +/- 1.0 mIU/mL (IU/L)], while the mean serum B-LH level as well as the LH B/I ratio was significantly lower in the impotent men throughout the observation period [11.4 +/- 2.0 (+/-SE) vs. 26.0 +/- 3.2 mIU/mL (IU/L) and 1.4 +/- 0.2 vs. 5.4 +/- 0.6; P less than 0.05 and P less than 0.02, respectively]. The B-LH pulse amplitude in the impotent men was reduced [mean peak LH, 8.6 +/- 0.3 vs. 25.3 +/- 4.0 mIU/mL (IU/L); P less than 0.05], while the LH pulse frequency was similar in the two groups. The median intrapulse LH B/I ratios were significantly higher than the median interpulse ratios in both impotent (P = 0.02) and normal men (P = 0.01).(ABSTRACT TRUNCATED AT 400 WORDS)     

Development of Junctions During Differentiation of Lens Fibers

Throughout the differentiation of eye lens epithelium into fibers, an extensive system of intercellular junctions develops. The junctional assembly is initially characterized by the accumulation of 9.0-nm intramembranous particles, forming linear rows in the matching plasma membranes of adjoining fibers. At the final stage of the fiber differentiation, the junctional particles are assembled in geometrically packed arrays. The formation of linear rows and bidimensional lattices of intramembranous particles probably favors reciprocal recognition of cell surfaces and specific cell-to-cell interlocking. Moreover, the existence of a rather rigid lipid core of the plasma membrane of eye lens fiber may promote the clustered distribution of intramembranous particles and facilitate the junctional assembly.     

Notch signaling is necessary but not sufficient for differentiation of dendritic cells

The Notch family of receptors plays an important role in regulation of cell differentiation via direct contact between hematopoietic progenitor cells (HPCs) and bone marrow stroma (BMS). However the precise contribution of Notch in dendritic cell (DC) differentiation is controversial. In 2 different experimental systems using Notch-1-null embryonic stem cells and Notch-1-deficient HPCs we have found that Notch-1 is necessary for DC differentiation. However, activation of Notch-1 and Notch-2 with cell-bound Notch ligand did not result in differentiation of mature DCs or macrophages. Instead, it caused accumulation of immature myeloid cells. Removal of feeder cells resulted in rapid differentiation of DCs and macrophages. Addition of interleukin 4 (IL-4) into the culture dramatically increased accumulation of functionally potent DCs. Lipopolysaccharide was not able to reproduce this effect. Thus, these data indicate that Notch signaling prevents differentiation of mature myeloid cells. Instead, it results in accumulation of precursors readily able to differentiate into mature DCs once the Notch signal is stopped (eg, after cell emigration from bone marrow) and in the presence of other additional differentiation signals provided by IL-4. Thus, Notch is required but not sufficient for DC differentiation.     

A Quantitative Approach to SIV Functional Latency in Brain Macrophages

Journal of Neuroimmune Pharmacology - Lentiviruses are retroviruses that primarily infect myeloid cells, leading to acute inflammatory infections in many tissues particularly, lung, joints and the...     

Self-Management: A Comprehensive Approach to Management of Chronic Conditions

For both clinical and economic reasons, the increasing number of persons living with chronic conditions represents a public health issue of growing importance. Emphasizing patient responsibility, and acting in concert with the provider community, self-management represents a promising strategy for treating chronic conditions—moving beyond education to teaching individuals to actively identify challenges and solve problems associated with their illness. Self-management also shows potential as an effective paradigm across the prevention spectrum (primary, secondary, and tertiary) by establishing a pattern for health early in life and providing strategies for mitigating illness and managing it in later life. We suggest ways to advance research methods and practical applications of self-management as steps in its future development and implementation.Improvements in health care have resulted in greater numbers of people living with multiple chronic conditions for longer periods of time. With this change, chronic illness is now a major focus of health care.1 At the same time, increased attention has been concentrated on approaches to manage chronic symptoms to maintain patient independence and quality of life over longer periods of time. Approaches to managing chronic illness are shifting from the traditional provider–patient relationship to a paradigm in which individuals with chronic conditions play a key role in guiding their care, in partnership with health care providers.2,3Many prevalent chronic conditions, such as heart disease, diabetes, and arthritis, though unique in their own attributes and demands, share common challenges associated with their management. These include dealing with symptoms and disability; monitoring physical indicators; managing complex medication regimens; maintaining proper levels of nutrition, diet, and exercise; adjusting to the psychological and social demands, including difficult lifestyle adjustments; and engaging in effective interactions with health care providers.4,5The identification and elaboration of common patient-centric strategies to deal with these challenges is the focus of the field of self-management.6,7 Regardless of the chronic condition, the development of a generic set of skills has proven successful in allowing individuals to effectively manage their illness and improve health outcomes.8 A 2010 report by the Department of Health and Human Services included self-management as one of 4 goals in a strategic framework for improving the health status of individuals with multiple chronic conditions.9 More recently, the 2012 Institute of Medicine report “Living Well With Chronic Illness: A Call for Public Health Action” included self-management10 as one of several models of living well interventions, noting that self-management programs instill individual responsibility and offer tools for patients to use in caring for their chronic illness.10There is increasing recognition that chronic illness, including its prevention, treatment and management, represents a public health as well as a clinical issue.11,12 Indeed, the Institute of Medicine report noted that a population health perspective for developing strategies, interventions, and policies to combat chronic illness is critical.10 Community-based self-management intervention programs are one aspect of a population-based approach addressing the larger public health problem of chronic conditions in the United States and across the globe. There is an extensive body of literature related to self-management of chronic conditions, but our intent with this article is not to provide a comprehensive review, but rather to highlight the unique contribution of nurse scientists to the field.Nursing science has enhanced the care of individual patients and has tested interventions that can be scaled up for implementation at the population level. We present examples of nursing science that demonstrate effectiveness, promise sustainability and scalability, and set the foundation for implementing wide-reaching public health actions for managing chronic illness.12There is increased awareness of the need to promote conceptual clarity regarding self-management and its integration into clinical practice. Equally important is the requirement to develop more sophisticated models of self-management, tailored to various health conditions and situations. Fundamental to the development of such models and their practical application is the need to conduct research that informs self-management practice and contributes to health policy.The nursing community, comprising both researchers and clinicians, plays a crucial role in efforts to provide the evidence base for innovative self-management practices, and is ideally positioned to implement those advances in a practical manner. Over the course of its history, the National Institute of Nursing Research (NINR) at the National Institutes of Health (NIH) has promoted self-management science as one of its core areas of investigation, supporting research to improve and manage symptoms of acute and chronic illness.13 Recognizing that self-management represents a topic of ever-increasing importance, a goal of NINR is to advance the science of self-management and, ultimately, disseminate results widely for translation into clinical practice.Recently, a group of nurse scientist leaders assembled to discuss “The Science of Chronic Illness Self-Management” as the topic of the 2013 National Nursing Research Roundtable. The Roundtable is an annual meeting with the purpose of providing a regular forum of communication about the direction and conduct of nursing research. Discussions from this year’s meeting resulted in a set of recommended areas of focus and approaches to advance the field and practice of self-management (see the box on this page).

Recommended Areas of Focus and Approaches to Advancing Self-Management