bluuebunny/crossref_metadata_2025 huggingface dataset:
Transmission electron microscopic demonstration of vimentin in rat osteoblast and osteocyte cell bodies and processes using the immunogold technique
Authors: Shapiro Frederic , Cahill Christopher , Malatantis George , Nayak Ramesh C.
Published: February 2005

Background: The immunogold labeling technique and transmission electron microscopy were used to demonstrate the expression and position of the intermediate filament vimentin in rat osteoblast and osteocyte cell bodies and cell processes. Conventional light and transmission electron microscopic studies of bone cells demonstrated adjacent cell linkage to be mediated by osteoblast and osteocyte processes present within the canalicular system traversing the bone matrix. The cell processes were filled with densely packed filaments, many of which have been shown previously to be actin microfilaments. The appearance, however, of 10 nm diameter filaments in some cell processes and the fact that the intermediate filament vimentin has been defined in many cells of mesenchymal origin raised the possibility that some of these filaments might be vimentin. The ultrastructural colloidal gold immunochemical technique allowed for demonstration in situ of the expression of vimentin filaments plus accurate definition of their position.

Methods: The studies were performed in newborn rat femoral and tibial diaphyseal cortical bone and in 1‐week‐old repair bone from 2.4 mm diameter defects made through the lateral cortex in 6‐week‐old rat femurs and tibias. The bone tissues for the immunochemical study were fixed in 1% glutaraldehyde, 4% paraformaldehyde, and 0.1 M phosphate buffer (pH 7.4) for 2 days. Decalcification was performed in 6% EDTA for 2–3 days. Infiltration involved use of Lowicryl resin K4M, and the embedding and curing processes were performed in a cryostat with temperatures −30°C. An antivimentin monoclonal antibody was used for labeling using the postembedding technique. Effective antibody dilutions ranged from 1:10 to 1:200, with the dilutions of 1:25 and 1:100 showing the best combination of filament labeling with the least matrix background. The grids were exposed to 10 nanometer gold colloid conjugated goat anti‐mouse IgM for demonstration of binding.

Results: Vimentin immunolabeling was defined clearly in relation to filaments within the osteoblast and osteocyte cell body cytoplasm, throughout the entire length of the osteoblast and osteocyte cell processes, and in close relationship to the intercellular gap junctions which were present within the cell processes both close to the cell bodies and within the canaliculi well away from them.

Conclusions: Immunogold labeling demonstrates the presence of the intermediate filament vimentin in osteoblast and osteocyte cell bodies and processes of rat bone. Vimentin distribution is not concentrated to specific areas, is present throughout the extent of the bodies and processes, and is seen immediately adjacent to gap junctions. © 1995 Wiley‐Liss, Inc.
Altered mitochondrial morphology of rat embryos in diabetic pregnancy
Authors: Yang Xiaolin , Håkan Borg L. A. , Eriksson UlF J.
Published: February 2005

Background: Previous studies in vivo and in vitro have suggeted that the oxidative metabolism of the embryo may have a role in the treatogenicity of diabetic pregnancy. In particular, the production of reactive oxygen species by the embroyonic mitochondiria has been implicated in the teratological process. The induction of congenital malformations by the diabetic milieu occurs during the early embryonic development. The present study aimed to estimate the role of the embryonic mitochondria in the teratological process of diabetic pregnancy by studying mitochondrial morphology in the embryos exposed to a diabetic environment in vivo or in vitro during early organogenesis and late fetal development.

Methods: For studies in vivo embryos of control or streptozotocin‐dia‐betic rats were taken at gestational days 9–11 and sujected to light and electron microscopical analysis. The brain, heart, and liver of day‐15 fetuses were also observed. For studies in vitro day‐9 embryos of normal rats were cultured in a whole‐embryo culture system for 48 hours. The culture media were supplied with high conectration of diabetes‐related substrates and metabolites, and their effect on structure of embryonic neuropeithelial cells determined.

Results: The light microscopoical observations demonstrated numberous cytoplasmic vaculoes in the ectoderm of day‐9 embryos and the neuroepithelium and blood cells of day‐10 and day‐11 embryos of diabetic rats. Ultrastructuraily, these vacuoles were found to be mitochondria undergoing large‐amplitude swelling with loss of matrix density and disturbed cristae. In contrast, no Mitochondrial differences were found in the brain, heart, and liver, when day‐15 fetuses from normal and diabeic rats were compared. Ultrastructural analysis of day‐9 embryos cultured for 48 hours in the presnce of high conectrations of D‐Glucose, pyruvate, β‐hydroxybutyrate, and α‐ketoisocaproate also showed high‐amplitude mitochondrial swelling in the neuroepithelium. The motochondrial swelling was, however, not found in embryos cultured in a high conectration of L‐glucose, excluding simple osmotic effects of the diabetes‐related substrated and metabolites.

Conclusions: The mitochondrial morphological changes appeared in embryos subjected to a diabetic environment during a time period when the congenital malformations in diabetic pregnancy are induced. The results support the notion that embryonic mitochondria are involved in the teratological process of diabetic pregnancy. © 1995 Wiley‐Liss, inc.
Estimation of capillary length density in skeletal muscle by unbiased sterological methods: II. Use of vertical slices of unknown thickness
Authors: Artacho‐Pérula Emilio , Roldán‐Villalobos Rafael
Published: February 2005

Background: The capillary network of the skeletal muscle was analyzed from a quantitative point of view with the purpose of determining the capillary length density—Lv(cap/mus). A recent stereological method was applied to estimate this quantity using vertical slice of unknown thickness.

Methods: First, the whole muscle was systematically sampled according to the fractionator method. The capillary length density was estimated on each chosen field of vision where the vertical axis was always identified as parallel to the major axis of the muscle fibers. Three measurements were performed: count of intersections between capillaries and cycloid test lines, count of intersections between capillaries and straight test lines, and count of capillary end points corresponding to the intersections of capillaries with the parallel planes of the vertical slices.

Results: The estimated capillary length density was 1,578 mm per mm3 of skeletal muscle tissue. The average thickness of the vertical slices was also estimated as 23.4 μm, which is roughly 6% less than the thickness measured using the microcator information on the microscope stage displacement.

Conclusions: The advantages of this methodology were bases on two main features: the method is assumption‐free on the degree of capillary and muscle anisotropy, and the thickness of the vertical slices need not be known nor constant. © 1995 Wiley‐Liss, Inc.
Skeletal alterations in hypophysectomized rats: I. A histomorphometric study on tibial cancellous bone
Authors: Yeh James K. , Chen Meng‐Meng , Aloia John F.
Published: February 2005

Background: Hypophysectomy (HX) results in a cessation of bone growth and a decrease in bone metabolism. The purpose of this study is to examine the effect of HX on the static and dynamic histomorphometry of cancellous bone in the secondary spongiosa of the proximal tibial metaphysis in rats.

Methods: Female rats, at 2 or 3 months of age, were HX and sacrificed at 0, 5 days, 2 and 5 weeks after the surgery. Age‐matched intact rats served as controls. Cancellous bone histomorphometry was performed on doublefluorescent labeled, 30‐um‐thick sections of the proximal tibia. Tartrateresistant acid phosphatase histomorphometry was performed at 5 days on HX and control rats to evaluate the resorption in the metaphyseal bone.

Results: Although the intact rats gained in body weight, tibial length, tibial weight, and density after 5 weeks, these changes did not occur following HX. As compared to the basal group, HX resulted in a decrease in the density and dry weight of the metaphysis. The histomorphometric data showed that the cancellous bone volume and trabecular number of the secondary spongiosa were decreased and the separation was increased in the HX rats. The dynamic results showed that HX significantly decreased longitudinal growth rate and tissue‐based bone formation and resorption. However, the bone surface‐based eroded surface, labeled surface, the mineral apposition rate, and the bone formation rate did not differ between the intact and the HX rats at either the 2 or 5 weeks study. Five days after HX, the bone surface and tissue‐based osteoclast surfaces were significantly lower in the HX than in the intact rats.

Conclusions: Pituitary hormone deficiency results in cancellous bone loss. The bone loss is due primarily to the suppression of longitudinal growth‐dependent bone gain and the inhibition of tissue‐based bone turnover with a lower bone formation relative to bone resorption. The surfacebased bone turnover is not affected. © 1995 Wiley‐Liss, Inc.
Skeletal alterations in hypophysectomized rats: II. A histomorphometric study on tibial cortical bone
Authors: Chen Meng‐Meng , Yeh James K. , Aloia John F.
Published: February 2005

Background: Pituitary hormones play an important role in bone growth, modeling, and remodeling. The purpose of this study is to examine the effect of hypophysectomy (HX) on tibial cortical bone with histomorphometry.

Methods: Forty‐Five female Sprague‐Dawiey rats, at 3 months of age, were hypophysectomized or served as intact controls. They were sacrificed at 0, 2, and 5 weeks after the surgery. Cortical bone histomorphometry was performed on double‐fluorescent‐labeled 30‐mcm‐thick sections of the tibial shaft.

Results: The dry weight and density of tibial diaphysis and the cortical bone area of the tibial shaft in the HX rats were significantly lower (P<0.05) than that of the age‐matched intact rats, but did not differ between the HX and basal control rats. The dynamic data show that the bone formation parameters (labeled surface, mineral apposition rate, and bone formation rate) were profoundly decreased (P<0.01) on both the periosteal and endocortical surfaces in the HX rats as compared with the age‐matched intact rats at the 2 and 5 weeks. However, the decrease in the labeled surface was much less on the endocortical envelope than on the periosteal envelope in the HX rats. Although no significant change was detected in the medullar size between the HX and age‐matched intact rats, the eroded surface on the endocortical surface was greater (P<0.05) in the HX rats than in the intact rats at either time point.

Conclusions: Hypophysectomy‐suppressed, radial growth‐dependent bone gain without a bone loss in the tibial shaft of the young rat. This is associated with decreased modeling‐dependent bone formation. A greater eroded surface on the endosteum did not affect the marrow size at 5 weeks after hypophysectomy. © 1995 Wiley‐Liss, Inc.
Neonatal hypothyroidism causes delayed sertoli cell maturation in rats treated with propylthiouracil: Evidence that the sertoli cell controls testis growth
Authors: de Franca Luiz Renato , Hess Rex A. , Cooke Paul S. , Russell Lonnie D.
Published: February 2005

Background: The testes of rats treated neonatally with propylthiouracil (PTU) grow to almost twice their normal size. The cause of testicular enlargement has been suggested to be the result of delayed maturation of Sertoli cells, allowing Sertoli cell division to occur beyond the 15th postnatal day, the commonly recognized cutoff date for Sertoli cell divisions. It has been shown that an increased population of Sertoli cells in postnatal development supports increased numbers of germ cells in adult animals. After examining developing rats treated neonatally with PTU, we hypothesized that an approximate 10‐day delay in maturation was occurring and proceeded to test this hypothesis experimentally. Thus the purpose of this report was to determine if a 10‐day delay in maturation could explain the increased numbers of Sertoli cells and increased testis size in PTU‐treated animals.

Methods: Both control animals and animals treated neonatally with PTU N = 5/group were sacrificed at 15 and 25 days of age and prepared for electron microscopy.

Results: Micrographs show and morphometric ultrastructural analysis of numerous parameters demonstrated at the 95% probability level that Sertoli cells from 25‐day‐old PTU animals are not different in size and most constituents (volume and surface area) from 15‐day‐old control animals and are less mature than 25‐day‐old control animals. Mitosis of Sertoli cells was observed in PTU‐treated animals in 25‐day‐old animals but not in agematched controls. The number of Sertoli cells in 25‐day‐old PTU‐treated animals is significantly increased over age‐matched controls. Micrographs show the presence of immature Sertoli cell nuclei in 25‐day‐old animals receiving PTU as well as increased germ cell degeneration in this group. Sertoli cell tight junction formation is also delayed in PTU‐treated animals as compared with controls.

Conclusions: Together, the data show that delayed maturation of Sertoli cells occurs in treated animals that corresponds to a minimum of 10 developmental days. In the immature state, Sertoli cells continue to divide. Data presented herein and published data related to PTU treatment indicate that delayed maturation of the Sertoli cell results in delayed maturation and proliferation of other testicular cell types. From this and from published data, the hypothesis is presented that the Sertoli cell is responsible for the overall control of testis development. © 1995 Wiley‐Liss, Inc.
Functional implication of autophagy in steroid‐secreting cells of the rat
Authors: Yi Jing , Tang Xue Ming
Published: February 2005

Background: Autophagy, while frequently observed in embryonic cells undergoing differentiation and in pathologically altered cells, appears to occur less commonly in normal, fully differentiated cells. Our previous work revealed that the frequency of autophagic activity was rather high in the Leydig cells of rat testes, but the functional significance of autophagy in Leydig cells remains obscure. The purpose of the present study is to investigate the possible role of autophagy in steroid‐secreting cells.

Methods: The autophagic activity was investigated in two steroid‐secreting cells, e.g., Leydig cells and adrenocortical fasciculata cells of rats. Cytidine monophosphatase (CMPase) cytochemistry was utilized to show the activity of lysosomal enzymes in autophagosomes. Electron microscopic morphometry was employed to analyze the frequencies of autophgy in the cells of the rats intact or treated with related hormones resulting in a hyper‐ or hypo‐secretion of testosterone and corticosterone.

Results: Autophagy took place in normal steroid‐secreting cells with higher frequencies than in many other cells including the tubular cells of kidney and hepatocytes. The large number of autophagosomes or autophagic vacuoles allowed to outline the autophagic process in these cells. The C‐shaped double‐membrane profiles tending to demacate a portion of cytoplasm were referred to as pre‐autophagosomes. So called early autophagosomes were the vacuoles enclosed completely by double delimiting membranes, containing normal‐looking cellular components. The majority of sequestered organelles appeared to be mitochondria and smooth endoplasmic reticulum. The autophagosomes starting digestion were considered as late autophagosomes or autophagic vacuoles, the indications of which were the destruction of their contents or the presence of lysosomal enzymes demonstrated by a positive CMPase reaction. Residual bodies were frequently observed to be exocytosed. The quantitative assay revealed an alteration of autophagic activity in close relation with steroid‐secreting states. The number of autophagosomes was one‐fold higher in hyposecreting Leydig cells after 2 days testosterone administration, and three‐fold higher in hyposecreting adrenocortical fasciculata cells after one dosage of dexamethasone administration. In addition, the autophagosomes showed a four‐fold decrease in hypesecreting Leydig cells stimulated by LRH for 2 days.

Conclusions: Considering that most of the autophagocytosed organelles were steroid‐producing apparatus, we may conclude that, by removing part of steroid‐producing organelles, autophagy might play a role in adapting to or even regulating the secretory activity. This hypothesis was strongly supported by the fact that the intensity of autophagy varied in company with the fluctuation of steroid secretion. © 1995 Wiley‐Liss, Inc.
Growth and turnover of rat otoconia as revealed by labeling with tetracycline
Authors: Kawamata Seiichi , Igarashi Yoshikazu
Published: February 2005

Background: The formation process of otoconia remains controversial, and the turnover rate of mammalian otoconia has not been determined.

Methods: Tetracycline was administered as a tracer for calcium, and the growth and turnover of rat otoconia were examined by fluoresence microscopy.

Results: Exposure of rats to tetracycline from 15.5 gestational day to 3‐day postpartum resulted in incorporation of the drug into central portions of all otoconia, in both maculae sacculi and utriculi. When postnatal rats were injected subcutaneously with tetracycline, uptake of the drug into otoconia depended on the age of rats at injection. Apparent fluorescence was emitted from the periphery of all otoconia when tetracycline was injected into 7‐day‐old or younger rats. However, very little or no fluorescence was observed when this reagent was administered 10 days after birth. No fluorescence was detected when rats of 12 days of age or older were given this antibiotic. Otoconia that had been labeled with tetracycline during gestation were monitored during subsequent development, and it was found that all otoconia retained labelling in their central portions for at least 12 months. No otoconia that were not labeled with tetracycline were found.

Conclusions: These findings indicate that: (1) all otoconia grow synchronously during late gestation and the neonatal period (up to about 10 days after birth) by accretion, (2) no new otoconia are formed subsequently, and (3) essentially no turnover of otoconia occurs and probably no turnover of calcium takes place under normal conditions once otoconia have formed. © 1995 Wiley‐Liss, Inc.
Relationships between endothelial cells, pericytes, and osteoblasts during bone formation in the sheep femur following implantation of tricalciumphosphate‐ceramic
Authors: Decker Brigitte , Bartels Helmut , Decker Sieghard
Published: February 2005

Background: The origin of osteoblasts is still controversial. Whereas several authors consider the stromal fibrablast of the bone marrow as the osteoprogenitor cell, others propose that the osteoblasts can be derived from the “capillary system.” The present study examines the replacement of tricalciumphosphate (TCP)‐ceramic implanted into an artificial bone defect by newly formed bone. The results support the hypothesis that osteogenic cells can be derived from invading blood vessels.

Methods: The spongiosa of the trochanter major of sheep was removed and the defect was filled with TCP‐ceramic. Two months after surgery the ceramic implants together with the surrounding bone were removed and processed for transmission electron microscopy. Serial ultrathin sections of three newly formed osteons were examined.

Results: The osteons contain one or two small sprouting capillaries, a peripheral layer of osteoblasts, and in between, a network of glycogen‐rich cells. Some of the glycogen‐rich cells are completely or partly surrounded by the endothelial basal lamina and are thereby characterized as pericytes. Weibel‐Palade bodies, which are considered to be a marker of endothelial cells, were occasionally observed in glycogen‐rich pericytes.

Conclusions: Since pericytes differentiate into osteoblasts under in vivo and in vitro conditions, and have thus been regarded as osteoprogenitor cells, the presence of Weibel‐Palade bodies in these cells suggest that osteogenic cells can be derived from endothelial cells. © 1995 Wiley‐Liss, Inc.
Three‐dimensional organization of the elastic fibres in the rat lung
Authors: Gonçalves Carlos Alberto , Figueiredo Maria Helena , Bairos Vasco António
Published: February 2005

Background: The elastic framework of the distal lung has been studied by light microscopy (LM) and transmission electron microscopy (TEM). The preservation of the elastic fibres, for the three‐dimensional observation in their relative positions, is difficult because they lack support when the normal methods of tissue processing are used. The goal of the present study was to understand the three‐dimensional ultrastructure and organization of the elastic fibres of the lung preserved in their relative positions.

Methods: A combination of intravascular resin injection and formic acid digestion was used. The resin cast of the microvasculature acted as a scaffold to preserve the in vivo arrangement of the elastic fibres that are, otherwise, easily collapsible. Scanning electron microscopy (SEM) samples were further processed for TEM in order to confirm that the fibres were indeed components of the elastic system.

Results: SEM demonstrated a fine framework of elastic fibres, representing remnants of the alveolar walls, with the casted capillaries interwoven with the network of elastin. Each individual elastic fibre is composed of a small bundle of discrete fibrils. Some of these fibrils emerge from the fibre and join other fibres, producing an anastomosing appearance. Several elastic fibres link the walls of the intrapulmonary conducting airways, the vessels walls and the alveolar network, thus establishing an interrelated and interlaced framework.

Conclusions: The method we have applied to visualize the elastic fibres of the lung is a unique approach to define the spatial organization of the pulmonary elastic fibres. We have demonstrated here the close relationship between the elastic fibres and the capillaries of the septal alveoli. The arrangement of the interwoven network of elastin and its relationship with the capillaries offers the structural setting for the distending capacity of the alveolar wall. © 1995 Wiley‐Liss, Inc.
Quantitative microscopic changes in the mole rat testes during an annual cycle
Authors: Gottreich Ahuva , Hammel Ilan , Yogev Leah , Terkel Joseph
Published: February 2005

Background: The mole rat Spalax ehrenbergi is a solitary, aggressive subterranean rodent. The present study summarizes a year‐round investigation of histological changes and sperm production in the mole rat testes.

Methods: The testes were processed for routine histological examination. Tissues were fixed in Bouins' solution embedded in paraffin blocks and stained by hematoxylin‐eosin. A systemic sampling approach was used to photomicrograph the tissues for histomorphometric assessment.

Results: Quantitative morphometry provided parameters indicating the activity of the tubuli testes during the year. The volume fraction of testes interstitial tissue from animals kept in captivity was unimodal and demonstrated an almost 3‐fold increase (from Vv = 0.19 ± 0.02 μm3/μm3 in October to 0.55 ± 0.05 μm3/μm3 in November) (i.e., early winter). The testes tubuli Vv of these animals demonstrated a significant decrease (from 0.75 ± 0.08 μm3/μm3 in October to 0.37 ± 0.04 μm3/μm3 in November).

The testes tubuli star‐volume demonstrated an annual bi‐modal distribution, during winter (November‐February, v* = 0.075 ± 0.015 μm3) and summer (July‐August, v* = 0.085 ± 0.016 μm3); the lowest star‐volume was found in April (v* = 0.017 ± 0.004 μm3 in the field group).

The testes tubuli revealed the entire sperm differntiation process throughout the year, from spermatogenesis (cells at all stages) to observation of mature spermatozoa, even though the mole rat is fertile only once a year during the winter. An increase in spermatogenesis and sperm production occurs during winter (Novermber‐December) and autumn (April‐May).

Conclusion: It would appear that although changes in spermatogenesis and tissue structure are found during the annual cycle, the males none the less have a fecundity potential throughout the entire year. ± 1995 Wiley‐Liss, Inc.
Reducing the levels of formaldehyde exposure in gross anatomy laboratories
Authors: Coleman Raymond
Published: February 2005

Background: A method is described in which formaldehyde levels are greatly reduced in our gross anatomy laboratory in order to comply with increasingly severe safety and health regulations.

Methods: A novel type of dissection “bed” has been introduced which incorporates an internal motor that causes a downflow of formaldehyderich vapors, which are absorbed by a replaceable active carbon filtration system.

Results: Use of the new dissection “beds” has resulted in the recirculated air being virtually formaldehyde‐free. Formaldehyde vapor levels in our gross anatomy laboratory have been greatly reduced and are typically in the range of 0.03–0.09 ppm.

Conclusions: The new system allows us to comply with safety and health regulations and provide a dissection room with an excellent working environment. © 1995 Wiley‐Liss, Inc.
Lipid‐lowering efficacy of 3,4‐di(OH)‐phenylpropionic L‐leucine in high‐cholesterol fed rats
Authors: Kim Soon‐Ja , Bok Song‐Hae , Lee SangKu , Lee Mi‐Kyung , Park Yong Bok , Kim Hye‐Jin , Choi Myung‐Sook
Published: February 2005

A preliminary study revealed that 3,4‐di(OH)‐hydrocinnamate (HC), a polyphenolic compound, lowered the plasma lipids in high‐cholesterol fed rats. Accordingly, this study was designed to test the lipid‐lowering efficacy of a synthetic derivative, 3,4‐di(OH)‐phenylpropionic (L‐leucine) amide (PPLA), in rats fed a high‐cholesterol (1%, wt/wt) diet. As such, HC or PPLA was given as supplement to a high‐cholesterol diet for 6 weeks at a dose of 0.137 mmol/100 g diet. The supplementation of HC and PPLA significantly lowered the plasma and hepatic cholesterol and triglyceride levels compared to the control group. The activities of hepatic HMG‐CoA reductase (164 ± 9.12 and 124.74 ± 17.09 pmol/min/mg protein vs. 245.41 ± 13.01 pmol/min/mg protein, p < 0.05) and ACAT (411.49 ± 11.48 and 334.35 ± 17.68 pmol/min/mg protein vs. 490.41 ± 16.69 pmol/min/mg protein, p < 0.05) were significantly lower in the HC‐ and PPLA‐supplemented groups than in the control group. However, PPLA was more effective in inhibiting the enzyme activities than HC. The excretion of neutral sterol was significantly higher in HC‐ and PPLA‐supplemented groups than in the control group. Therefore, these results indicate that PPLA, a leucine‐attached version of HC, exhibited a similar significant hypocholesterolemic effect to HC in rats fed a high‐cholesterol diet. © 2005 Wiley Periodicals, Inc. J Biochem Mol Toxicol 19:25–31, 2005; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.20054
Pattern of CO2 fixation during vegetative development and gametic differentiation in Chlamydomonas reinhardtii
Authors: Kates Joseph R. , Jones Raymond F.
Published: February 2005

The rate of C14O2 incorporation into amino acids and organic acids in C. reinhardtii is a function of particular stages of development in the life cycle of the alga. Gametic differentiation in nitrogen free medium is accompanied by a reduced rate of amino acid synthesis and a higher synthesis of organic acids than that found for the cells undergoing vegetative development. The addition of ammonium to differentiating gametes results in an increased synthesis of amino acids, particularly the basic ones, and a concomitant reduction in organic acid synthesis.
Heat production during diving in the fresh water turtle, Pseudemys scripta
Authors: Jackson Donald C. , Schmidt‐Nielsen Knut
Published: February 2005

Heat production was measured calorimetrically in the turtle, Pseudemys scripta elegans, (a) while submerged and (b) while partially submerged and breathing various gas mixtures. Submersion resulted in a profound reduction of heat production (80%). This reduction was not merely a response to the dive per se, but depended on the oxygen concentration available to the turtle prior to the onset of the dive. Heat production while breathing gas mixtures with different O2 concentrations was unchanged down to 5% O2. At 3% O2, heat production was 50% of normal, and at 100% N2, it was 20% of normal. Uptake of dissolved O2 from water was found to be 6% of the O2 uptake from air by these turtles. These results suggest that following diving, there is a profound reduction in metabolic rate, but not until the O2 stores are depleted. This low rate is primarily anaerobic and only a very low oxidative metabolism can be supported by O2 extracted from the water.
The effect of hematoporphyrin and light on human platelets. I. Morphologic, functional, and biochemical changes
Authors: Zieve Philip D. , Solomon Harvey M. , Krevans Julius R.
Published: February 2005

The morphologic, functional, and biochemical changes produced by hematoporphyrin and light in human platelets have been characterized. by phase microscopy the cells appeared swollen and resembled signet rings; by electron microscopy they showed considerable loss of cytoplasm and their contour was smoother than normal. irradiated platelets were not aggregable by thrombin and calcium chloride, although they contained clottable protein, and were incapable of supporting clot retraction. a linear relationship was demonstrated between the per cent depletion of serotonin from irradiated platelets and the log dose of hematoporphyrin. the depletion of serotonin from these platelets was related lineraly to the log of time of exposure to light during the initial six minutes of exposure; but thereafter continued at a constant rate. the temperature of incubation influenced directly the rate of depletion of serotonin from irradiated platelets but did not influence the movement of serotonin into these platelets. atp was diminished considerably in irradiated platelets. these changes are attributable to damage to the membrane of the platelet by hematoporphyrin and light.

These studies provide additional information about the blood platelet in terms of its response to photodynamic action.
Circadian activity rhythms in cockroaches. III. The role of endocrine and neural factors
Authors: Def. Roberts Shepherd K.
Published: February 2005

The control of circadian activity rhythms (diurnal rhythms) in insects has been suggested to result by periodic neuroendocrine secretions. More specifically, Harker ('56) claimed that the locomotor rhythm in the cockroach, Periplaneta americana, is timed by a secretory “clock” located in the subesophageal ganglion. Later experiments by Harker indicated that this “clock” function failed unless the retrocerebral organs were left intact; allatectomy was said (no evidence given) to abolish a rhythm. The procedure for demonstrating a “clock” function in the ganglion involved transplanting it from a rhythmic donor into the hemocoel of an arrhythmic host and observing that the host subsequently became rhythmic. This result (without explicit information about the phase of the rhythm) does not warrant the conclusion that the ganglion acts as a clock. Therefore, I have attempted to confirm and extend these important results. Employing techniques essentially identical to Harker's, and using the same species of roach, I have been unable to find any evidence to support the original claim: (1) in 20 test animals, implantation of ganglia from rhythmic donors failed to re‐instate a rhythm, and (2) allatectomy (22 cases) or removal of the entire retrocerebral complex (20 cases) did not interfere with the rhythm.

The results of another series of experiments show that the cockroach brain is involved in the control of the activity rhythm. When the brain is surgically bisected (mid‐sagittal) through the pars intercerebralis, arrhythmic activity patterns are immediately evoked. These continue for many weeks, but in a few cases rhythms ultimately “regenerate”.
Purine pyrophosphorylase as a selective genetic marker in a mouse lymphoma, P388, in cell culture
Authors: Ozer Harvey L.
Published: February 2005

It is possible to use the purine pyrophosphorylase in mammalian cell culture systems as a genetic marker in selecting small numbers of enzyme positive cells from large populations of pyrophosphorylase negative cells of the mouse lymphoma line P388 in medium containing amethopterin, hypoxanthine, glycine and thymidine. Conversely, it is readily possible to obtain pyrophosphorylase‐deficient cells by treatment with 8‐Azaguanine. We were unsuccessful in demonstrating DNA‐mediated transformation using DNA from enzyme positive cells incubated with cells which were enzyme negative.
Lysosomes in Tetrahymena pyriformis I. Some properties and lysosomal localization of acid hydrolases
Authors: Müller Miklós , Baudhuin Pierre , De Duve Christian
Published: February 2005

In homogenates of Tetrahymena pyriformis, five hydrolases — phosphatase, ribonuclease, deoxyribonuclease, proteinase, amylase — with acid pH optima were found. Over 75% of their activity is sedimentable with a centrifugal force of 250,000 g. min. Only 17% of the acid phosphatase and ribonuclease is active when assayed in the presence of 0.25 M sucrose at 0°. Exposure to a lowered osmotic pressure, freezing and thawing, and incubation at temperatures over 0° result in activation of the latent phosphatase and ribonuclease. After isopycnic centrifugation in a sucrose density gradient the hydrolases show a broad distribution which differs greatly from those of enzymes associated with mitochondria (succinate dehydrogenase) or with peroxisomes (catalase). The results are interpreted as evidence that the five acid hydrolases studied are localized in lysosomes which represent a distinct population of subcellular particles in Tetrahymena.
The cloning of macrophages in agar: Identification of the cells by electron microscopy
Authors: Lagunoff David , Pluznik Dov H. , Sachs Leo
Published: February 2005

A normal cell type that can be cloned in agar has been identified by electron microscopy as a macrophage.
Studies on carcinogenesis by avian sarcoma viruses. V. Requirement for new DNA synthesis and for cell division
Authors: Temin Howard M.
Published: February 2005

Partially synchronized secondary cultures of chicken embryo fibroblasts were exposed to Rous sarcoma virus (RSV) at various times in the cell cycle. The initiation of normal virus production could be inhibited by treatment with excess thymidine or with cytosine arabinoside without any effect on mitosis. This result is consistent with the hypothesis that the provirus of RSV is DNA, although the virion of RSV contains only RNA. It was, also, found that treatment which prevented or interferred with the first mitosis after exposure to virus prevented the initiation of normal virus production. Later mitoses did not appear to be necessary. A corollary of this finding is that virus production is synchronized by mitosis and that the length of the latent period depends upon when in the cell cycle a cell is exposed to virus.
A cytological study of the capacity for differentiation of normal hemopoietic colony‐forming cells
Authors: Wu A. M. , Till J. E. , Siminovitch L. , McCulloch E. A.
Published: February 2005

A two‐stage procedure has been used to obtain hemopoietic spleen colonies derived from single precursor cells containing radiation‐induced chromosomal markers. Of a total of 46 colonies examined, 17 were found to contain cells with abnormal karyotypes. In each of the 17 marked colonies, 90% or more of the dividing cells in the colony carried the same marker. Cell suspensions prepared from each of the individual colonies were tested for their content of dividing cells possessing recognizable differentiated functions. Metaphase cells with peroxidase‐positive granules in their cytoplasm were considered to be members of the granulopoietic series, while metaphase cells which contained Fe55 were considered to be members of the erythropoietic series. Results were obtained for 12 of the marked colonies, and in nine of these, the percentage of metaphases lacking the marker was less than the percentage of metaphases which were scored as erythropoietic, and also was less than the percentage of metaphases scored as granulopoietic. This is the result which would be expected if the marker were present in both erythropoietic and granulopoietic cells. These results provide support for the view that colony forming hemopoietic stem cells are multipotent, and that differentiation along more than one pathway can occur during the formation of macroscopic splenic colonies.
Variation in fructose transport among beef erythrocytes
Authors: Laris Philip C.
Published: February 2005

Erythrocytes from different cattle were shown to fall into two distinct groups with respect to fructose transport. The rates of entrance of fructose into the two classes of cells differed by a factor of 5 to 10. It was also found that one pathway, the slower, was susceptible to inhibition by glucose while the other was not. The characteristics of the two fructose transport systems remained constant for as long as a week (longest time studied) upon storage at 2°C. The differences observed could not be attributed to handling of the blood, to the relative glucose permeability, or to the age or sex of the animal. This work indicates that two different fructose transport systems are operating in beef erythrocytes of different individuals.
Delay in the cleavage of Tetrahymena pyriformis exposed to high hydrostatic pressure
Authors: MacDonald A. G.
Published: February 2005

Early cleavage stages of Tetrahymena pyriformis were exposed to 250 atms pressure for periods ranging from 20–200 minutes. Pressure halted cleavage which, in those cells exposed for only a short time, was resumed soon after decompression. Cells exposed to longer periods of pressure treatment failed to resume cleavage soon after decompression but cleaved after a delay approximating to a generation time. In the former case, the cleavage furrow remained intact; whereas in the latter case, the cleavage furrow frequently resorbed and a new furrow was formed to accomplish the delayed cleavage.

The results appear consistent with the hypothesis that the hypothetical macro‐molecular entity P2 of Rasmussen and Zeuthen is pressure labile and involved in the mechanical process of cleavage.
Application of irreversible thermodynamics to a functional description of the tumor cell membrane
Authors: Hempling H. G.
Published: February 2005

The permeability of the membranes of several strains of Ehrlich mouse ascites tumor cells to a series of homologous glycols was analyzed with equations from irreversible thermodynamics. Experimental data were fitted to the appropriate equations of Kedem and Katchalsky by means of an analog computer and values for σ, Lp, and ωRT were calculated. These parameters would become part of a functional profile of the membrane of the tumor cell. The glycols cross the membrane with energies of activation ranging from 16 kcal/mole for ethylene glycol to 20 kcal/mole for triethylene glycol. Entropies of activation ranged from 17 to 27 entropy units. This analysis provided several additional dividends. It is one of the few examples with a mammalian cell where the kinetics of non‐electrolyte transport have been described completely by the equations of irreversible thermodynamics. It has provided the means to distinguish how much of the transport occurred through porous paths in the membrane and how much involved solution within the matrix of the membrane. Finally, several tumor cell populations have been distinguished through these functional properties of their membranes.
Cytidine aminohydrolase activity in intact cultured transformed cells
Authors: Ellem K. A. O.
Published: February 2005

Cultures of several cell lines convert cytidine, if present in their medium, to uridine. The reaction is rapid, being virtually complete within one hour. The enzymatic activity is that of cytidine aminohydrolase (EC 3.5.4.5). The activity is exhibited by the intact cell — substrate and products being found in the medium bathing the cells. The activity should be taken into account in studies involving cytidine, and other 6‐amino pyrimidine nuclecsides.

Of nine transformed cell lines, and nine primary and secondary cell strains screened for the presence of cytidine aminohydrolase activity, six cultures were positive. All the positive cultures were heteroploid transformed lines. No diploid strain cultures were positive, and two near diploid mouse lymphoid neoplasms were negative. PPLO contamination was not detected in the positive cultures.

Within the limited series of cell cultures screened in this study, there appears to be a correlation between heteroploidy and the ability of the intact cultures to deaminate cytidine in excess of the cell requirement for the precursor.
Production of heparin related glycosaminoglycans by an established mammalian cell line
Authors: Kraemer Paul M.
Published: February 2005

A sulfated glycosaminoglycan has been isolated from the acid‐soluble fraction of an established line of Chinese hamster fibroblasts grown in suspension culture. This material has a molecular weight between 5000 and 10,000, contains equimolar amounts of hexosamine and uronic acid (orcinol method), and about 0.6 sulfate groups per hexosamine residue. About 80% of the sulfate groups are N‐sulfates on the basis of lability of the sulfate and the formation of equivalent numbers of free amino groups upon mild acid hydrolysis. The material is completely resistant to testicular hyaluronidase but is degraded to reducing monosaccharides and small oligosaccharides upon treatment with lyophilized cells of Flavobacterium heparinum that were grown on heparin. It is thought, therefore, to be related to the known N‐sulfated glycosaminoglycans heparin and heparitin sulfate.
Differentiation of antibody forming cells in lymph nodes during the anamnestic response
Authors: Schoenberg M. D. , Moore R. D. , Stavitsky A. B. , Gusdon J. P.
Published: February 2005

The cellular and subcellular events in the anamnestic response were considered. Rabbits previously immunized with key hole limpet hemocyanin (KLH) were given an anamnestic challenge in the hind footpads. The popliteal lymph nodes were removed at intervals after immunization and the following correlated on a temporal basis: the changes in the number and types of cells in the lymph nodes; the formation and regression of ribosomes, polyribosomes, endoplasmic reticulum and Golgi apparatus in plasma cells; the changes in intracellular immunofluorescence for anti‐hemocyanin; and, the incorporation of 14C labeled amino acids by lymph node cells into anti‐KLH during a brief in vitro culture period.

Maximum intracellular fluorescence for anti‐KLH and the largest incorporation of 14C labeled amino acids into antibody occurred between the third and fourth day after immunization. During this interval highly differentiated plasma cells were most numerous with respect to the total cellular population. These events took place in a 12 to 24 hour period.

This was followed by an abrupt decline in the synthesis of antibody. Coincident with this was a reduction in the number of recognizable plasma cells in the nodes, diminished intracellular fluorescence for anti‐KLH and a simplification of the cytoplasm of the plasma cells toward a lymphocytic form.
A study of the intracellular transport of calcium in rat heart
Authors: Patriarca P. , Carafoli E.
Published: February 2005

The distribution of in vivo injected 45Ca++ in the subcellular fractions of rat heart has been studied. Most of the radioactivity of the cell was found to be associated with the subcellular organelles; only a small fraction was recovered in the soluble phase. Mitochondria contained the greatest part of the total radioactivity associated with the subcellular organelles. After injection of 45Ca++ the specific activity of the mitochondrial calcium pool was several times higher than that of the calcium of the sarcoplasmic reticulum. Pentachlorophenol has been administered to rats to uncouple oxidative phosphorylation in heart mitochondria in vivo and its effect on the distribution of 45Ca++ in the heart studied. Under these conditions, it has been found that mitochondria contained much less 45Ca++ than the controls; this decrease was paralleled by an increase of the radioactivity associated with the microsomes and with the final supernatant. Experiments in which 45Ca++ was added to heart homogenates at 0° indicated that 45Ca++ also became bound to mitochondria and the other subcellular structures at 0°. However, PCP had no effect on the distribution of radioactivity among the subcellular fractions under these conditions. The results suggest that (1) energy‐linked movements of Ca++ take place in mitochondria of the intact rat heart, (2) a part of the uptake of 45Ca++ by mitochondria does not depend on metabolism, and, (3) the movements of Ca++ in heart mitochondria in vivo are probably more active than those in the sarcoplasmic reticulum.
Specificity of carotenoid transfer in the larval medaka, Oryzias latipes
Authors: Takeuchi Kunisuke
Published: February 2005

Carotenoids were introduced into the egg yolk of the medaka by either injection or feeding. Carotenoids for injection were dissolved in either olive oil, a mixture of olive oil and castor oil or Tween 80. Capsanthin, lutein and canthaxanthin were readily transferred from the yolk to the larval xanthophores, but no β‐apo‐8′‐carotenal and little β‐carotene were transferred.
The effect of DMSO on the permeation of non‐electrolytes through the barnacle cell membrane
Authors: Bunch Wilton H.
Published: February 2005

Frequent reports have suggested that Dimethylsulfoxide (DMSO) increases the flux of other molecules through biological membranes. This paper reports experiments in the single barnacle cell which permits differentiation of trans‐membrane fluxes from those utilizing intercellular pathways. Several non‐electrolytes were injected and wash‐out rates observed. There was no change in the time course of the wash‐out of these molecules when DMSO was added to the injected fluid. The conclusion of these experiments is that DMSO in low concentration does not change the permeation of non‐electrolyte across the cell membrane.
Cellular uptake of 3H‐actinomycin D in the hematological tissues and liver of the mouse
Authors: Cohen Natalie S.
Published: February 2005

Actinomycin D(AM), an inhibitor of DNA‐dependent RNA synthesis, produces a reversible cessation of red blood cell production. This study examines the in vivo cellular uptake of 3H‐AM in the hematological tissues and livers of B6D2F1 mice. 3H‐Am (sp. act. = 2.97 to 4.20 C/mmole) was given IV at a dose of 4.0 to 5.7 μg (14 μc) per mouse. Spleen, bone marrow, blood, and liver samples were taken for autoradiography at post‐injection times of five minutes to 67 hours.

We have confirmed the rapid in vivo cellular uptake of AM; substantial quantities of the drug were in the nuclei within five minutes of IV administration.

Not all cell types became labeled. Erythroid, hepatic, lymphoid, and reticulo‐endothelial (RE) cells and monocytes took up the label, whereas labeling of granulocytic elements was doubtful.

Most heavily labeled were liver cells (highest mean grain count = 110.1) and splenic RE(19.1) and erythroid (16.1) cells. Erythroid cells in the spleen were more heavily and more rapidly labeled than those in the bone marrow.

All nucleated erythroid maturational stages, in both the spleen and the bone marrow, were labeled, even at five minutes.

The time course of erythroid and hepatic labeling was quite different. Whereas early erythroid cells required six hours to become 100% labeled, liver cells were 100% labeled at five minutes and loss of hepatic labeling began as early as 15 to 30 minutes.