The hagfish slime gland thread cell. I. A unique cellular system for the study of intermediate filaments and intermediate filament-microtubule interactions.

Thread cell differentiation in the slime gland of the Pacific hagfish Eptatretus stouti has been studied using light microscopy and scanning and transmission electron microscopy. Thread cell differentiation is remarkable in that the life history of the cell is largely dedicated to the production of a single, tapered, cylindrical, highly coiled, and precisely packaged cytoplasmic thread that may attain lengths of 60 cm and diameters approaching 1.5 micron. Each tapered thread, in turn, is comprised almost entirely of large numbers of intermediate filaments (IFs) bundled in parallel. During differentiation of the thread, the IFs become progressively more tightly packed. Various numbers of microtubules (MTs) are found among the bundled IFs during differentiation of the thread but disappear during the latter stages of thread differentiation. Observations of regularly spaced dots in longitudinal bisections of developing threads, diagonal striations in tangential sections of developing threads, and circumferentially oriented, filament-like structures observed at the periphery of developing threads cut in cross section have led us to postulate a helically oriented component(s) wrapped around the periphery of the developing thread. The enormous size of the fully differentiated thread cell, its apparent singular dedication to the production of IFs, the ease of isolating and purifying the threads and IF subunits (see accompanying paper), and the unique position of the hagfish in the phylogenetic scheme of vertebrate evolution all contribute to the attractiveness of the hagfish slime gland thread cell as a potential model system for studying IF subunit synthesis, IF formation from IF subunits, aggregation of IFs into IF bundles and the interaction(s) of IFs and MTs.

Hagfish slime gland thread cells. II. Isolation and characterization of intermediate filament components associated with the thread.

The slime glands of hagfish have two major cell types, gland thread cells (GTCs) and gland mucous cells (GMCs), both of which upon contact with water contribute to the formation of an abundant quantity of viscous mucus. In previous studies we reported a method for the isolation of GTCs and showed that each ellipsoidal thread cell normally contains a single tapered thread which is uniquely coiled into a space-saving conformation and occupies most of the cell volume. Subsequently, the developing thread was found to consist mainly of intermediate filaments (IFs) aligned in parallel not only to one another but also to a far fewer number of interspersed microtubules (see accompanying paper). In the present report, urea extracts of GTCs were purified and characterized to establish the properties of thread components. One major (alpha) and two minor (beta, gamma) components prepared by anion exchange chromatography were shown to have similar apparent molecular weights of 63,500 +/- 500 daltons but different isoelectric pH values (alpha, 7.56; beta, 5.67; gamma, 5.31). Although the amino acid content of alpha differed significantly from beta and gamma, each of the three was highest in Gly, relatively high in Glx, Ser, Thr, Asx, Ala, Val, and Leu, and relatively low in Cys/2 and Trp. The amino acid compositions of beta and gamma were very similar, and only beta showed evidence of carbohydrate. The threonine content of the alpha component was higher than has been reported for IFs of different origin, and the high content of hydroxyamino acids (18, 19 residues per 100) in alpha, beta, and gamma has been approached only by several IF polypeptides from human or bovine epidermal keratins. Mixtures of the purified components formed 9-11-nm filaments in vitro. The results indicate that the hagfish thread cell is a rich source of IFs, which have a structure that facilitates formation of macrofibrils within the cell.

The hagfish slime gland: a model system for studying the biology of mucus.

The hagfish slime gland may provide a model system for studying certain aspects of the biology of mucus. Mucus is obtained in nonhydrated form by electrically stimulating the anesthetized hagfish and the secretions are stirred into ammonium sulfate. Centrifugation and filtration are than used to isolate the two major secretory products, mucous vesicles and threads. Specific advantages of the model and potential applications for research are discussed.

Threads in the hagfish slime gland thread cells: organization, biochemical features, and length.

Scanning electron microscopy in conjunction with cell isolation procedures revealed details of the packing of threads in hagfish slime gland thread cells. Biochemical studies indicate that the thread is largely composed of a protein subunit with a molecular weight of 63,500. Mathematical calculations suggest that the thread may attain lengths of 60 centimeters or more.

Adriamycin-induced inhibition of melanoma cell invasion is correlated with decreases in tumor cell motility and increases in focal contact formation.

Tumor cell adhesion to the extracellular matrix (ECM) is closely linked with tumor cell invasion and metastasis. In this study, we demonstrate that low levels of adriamycin, a widely used anticancer drug, can inhibit the invasion of highly metastatic K1735-M2 mouse melanoma cells in vitro through a reconstituted basement membrane extract. Adriamycin-induced inhibition of melanoma cell invasion occurred at levels of the drug (i.e. 1 ng/ml) that did not inhibit tumor cell growth, suggesting that the observed inhibition in tumor cell invasion was not due to the well-documented ability of adriamycin to interfere with DNA and/or RNA synthesis. Rather, these studies indicated that adriamycin-induced inhibition of melanoma cell invasion was accompanied by a corresponding decrease in the ability of adriamycin-treated tumor cells to migrate in response to several isolated ECM components including fibronectin, laminin and basement membrane (type IV) collagen. The decreased migration of adriamycin-treated tumor cells was not accompanied by a decrease in the adhesion or spreading of the adriamycin-treated cells on substrata coated with these ECM components. Instead, adriamycin-treated cells actually exhibited a slightly increased propensity (compared to untreated control cells) to adhere on fibronectin-, laminin-, and type IV collagen-coated substrata. Additionally, adriamycin treatment caused a dramatic increase in focal contact formation by these melanoma cells, as assessed by fluorescent microscopy of actin and vinculin. In addition to providing a useful model for which to study the molecular and cellular basis for focal contact formation, these results further emphasize the results of several other investigators that have suggested an important role for focal contacts in modulating tumor cell motility, invasion and metastasis.

Repair of left ventricular aneurysm. Changes in ventricular mechanics, hemodynamics, and oxygen consumption.

Anteroapical left ventricular aneurysms were produced in 23 sheep by coronary arterial ligation. Plication of the aneurysm does not change stroke volume or cardiac output and does not significantly change left ventricular oxygen consumption from the preoperative value of 5.1 +/- 2.6 ml/100 gm per minute. Plication, however, does increase left ventricular end-systolic elastance from 3.2 +/- 0.9 to 4.4 +/- 1.5 mm Hg/mm (p = 0.005). In nine of these sheep the midsagittal plane of the left ventricle was imaged by means of an array of sonomicrometry crystals before and after plication of the aneurysm. Regional wall stresses at end-systole and end-diastole and changes in diastolic function were calculated for anterior and posterior ventricular walls in the border zone adjacent to the aneurysm and in more basilar myocardium remote from the infarct. Plication significantly reduced end-systolic wall stresses and systolic stress integrals in the posterior border zone and remote myocardium, but it did not significantly change anterior wall systolic stresses or stress integrals. Plication also decreased diastolic stretching of border zone myocardium. Plication of anteroapical left ventricular aneurysm produced a shorter, more spherical ventricle and removed the dyskinetic segments but altered deformation (strain) in both circumferential and longitudinal directions. The changes in ventricular wall geometry and deformation provide an explanation for the increased ventricular end-systolic elastance and unchanged stroke volume observed after aneurysm plication.

The stretched ventricle. Myocardial creep and contractile dysfunction after acute nonischemic ventricular distention.

The hypothesis that nonischemic distention of the arrested, flaccid ventricle causes myocardial creep and reduces ventricular contractile force was tested in 16 sheep. Left ventricular volume was calculated from ultrasonic dimension transducers spanning left ventricular major and minor axes and left ventricular wall thickness. Changes in left ventricular volume were plotted against left ventricular pressure, with and without temporary occlusion of both venae cavae before and after nonischemic distention of the continuously perfused, flaccid nonbeating left ventricle arrested with oxygenated, normothermic blood-potassium perfusate. During 12 minutes of cardiac arrest, an apical balloon progressively distended the left ventricle to a peak pressure of 40 mm Hg in 11 sheep using a protocol designed to prevent subendocardial ischemia or mechanical injury. Coronary sinus lactate measurements and myocardial distribution of microspheres confirmed the absence of ischemia in 16 animals. In five control sheep the balloon was inserted but not inflated. Left ventricular volume at zero pressure increased from 5.9 +/- 3.5 to 9.5 +/- 4.4 ml (p < 0.05) after balloon inflation and did not change in the control animals. After maximum distention of the balloon, static left ventricular volumes at identical pressures were significantly greater. After passive distention, the slope of the end-systolic pressure-volume relationship, a measure of contractility, decreased significantly (p < 0.05) from 7.1 +/- 2.8 to 3.5 +/- 1.8 mm Hg/ml and did not change in the control group. Passive distention ("stretching") of the nonischemic flaccid left ventricle thus causes myocardial creep and reduces ventricular contractility.

Heparinless partial cardiopulmonary bypass for the repair of aortic trauma.

OBJECTIVE: We hypothesized that partial cardiopulmonary bypass with a heparin-bonded system would be a technically simple, effective adjunct for reducing paraplegia during repair of traumatic aortic rupture. It avoids the risk of heparin, but, unlike left atrial-arterial bypass, it can heat, cool, oxygenate, and rapidly infuse volume if needed. METHODS: A retrospective review was conducted of patients admitted for aortic trauma from July 1994 to December 1999. Bypass consisted of femoral venous (right atrial) cannulation, a centrifugal pump, and an oxygenator-heater/cooler. Arterial return was to the femoral artery or distal aorta. The entire system was heparin-bonded and no systemic heparin was given. RESULTS: Heparin-bonded partial bypass was established in 50 patients (mean age 43 +/- 17 years). Crossclamp time was 32 +/- 11 minutes (range 14-70 minutes), mean flow 3.0 +/- 0.8 L/min, and bypass time 64 +/- 43 minutes. During repair, 58% of patients received volume through the system (mean 1.1 +/- 1.9 L). Core temperature rose slightly (35.9 degrees C +/- 0.7 degrees C to 36.3 degrees C +/- 0.8 degrees C). Three of the 15 patients who underwent percutaneous femoral arterial and venous cannulation concomitant with their angiograms had vessel injury, with one limb loss, and this procedure was discontinued. Thirty-five patients underwent percutaneous femoral vein and direct distal aortic cannulation without event. The mortality rate for patients supported by bypass was 10%, and all deaths were due to other injuries. There were no new cases of paraplegia and no worsening of intracranial or pulmonary injuries. CONCLUSIONS: Heparin-bonded bypass is technically simple to use and avoids the risk of anticoagulation. Paraplegia was avoided. The ability to correct hypothermia, oxygenate, and rapidly infuse volume may simplify management and improve outcomes.

Pathogenesis of ischemic mitral insufficiency.

We developed a new animal model of ischemic mitral insufficiency in sheep and used it to test the hypothesis that the combination of posterior papillary muscle infarction and left ventricular dilatation was required to produce mitral regurgitation after acute inferior myocardial infarction of moderate size. In 12 sheep, ligation of the first two circumflex marginal coronary arteries infarcted 23% of the left ventricular mass, increased left ventricular cavitary area from 13.2 +/- 1.2 cm2 to 20.0 +/- 2.7 cm2 by 8 weeks and did not produce ischemic mitral regurgitation. In 13 sheep, ligation of the second and third circumflex marginal arteries infarcted 21% of the left ventricular mass and, in 11 of these sheep, the posterior papillary muscular mass as well. When the papillary muscle was included, this infarction produced progressively severe mitral regurgitation over 8 weeks, as left ventricular cavitary area increased from 12.5 +/- 2.6 cm2 to 22.8 +/- 3.8 cm2. We conclude that neither posterior papillary muscle infarction nor left ventricular dilatation alone produces ischemic mitral regurgitation after moderate-sized inferior wall infarction, but that the combination does.

Release of vasoactive substances during cardiopulmonary bypass.

Cardiopulmonary bypass is associated with bleeding and thrombotic complications, massive fluid shifts, and cellular and hormonal defense reactions that are collectively termed "the whole body inflammatory response." A host of vasoactive substances are produced, released or altered during cardiopulmonary bypass. These hormones, autacoids, and cytokines react with specific receptor proteins distributed throughout the body, and mediate the vascular smooth muscle and endothelial cell contractions that are responsible for much of the morbidity associated with open heart operations. This essay briefly reviews the actions, sources, and perturbations of the approximately 25 vasoactive substances known or believed to be altered by cardiopulmonary bypass, and provides an introductory reference list.

Left ventricular assistance without thoracotomy: mediastinal and transseptal approaches to the left heart.

Two methods to cannulate the left atrium for initiating mechanical left ventricular circulatory assistance using a centrifugal pump were investigated in 25 sheep. A modified Dennis transatrial septal approach produced flow rates of 88.6 +/- 14 mL.kg-1.min-1 through 21F catheters inserted during fluoroscopy through the jugular vein. In 8 animals the septal perforation was plugged after decannulation with a modified Rashkind umbrella plug. Fibroendothelial tissue covered the plug by 4 week. In 7 other animals, the septal defect was not plugged. The septal defect reached pinpoint size by 2 weeks and was completely closed by 4 weeks. In 10 sheep, the left atrium was cannulated from the neck through the mediastinum. Left ventricular assistance flow averaged 71.6 +/- 14 mL.kg-1.min-1. Mean blood loss during 1 hour of left ventricular assistance was 47 mL. In 8 animals, the atrial perforation was plugged with a mean blood loss of 253 +/- 194 mL. In 2 animals, the perforation was intentionally not plugged; mean blood loss was 700 mL. All animals survived. The modified Dennis transatrial method is recommended as a safe, expeditious, cost-effective method to implement left ventricular assistance without thoracotomy. The mediastinal approach, which is technically possible in humans, is more difficult but feasible. Left ventricular assistance has been proven to be the most effective way to rest the failing, ejecting left ventricle. Implementation without thoracotomy potentially expands applications of left ventricular assistance for temporary support of patients with severe manifestations of ischemic heart disease.

Traumatic aortic rupture: recent outcome with regard to neurologic deficit.

BACKGROUND: Traumatic aortic rupture is highly lethal, and its surgical treatment is complicated by a high rate of paraplegia. METHODS: The charts of 263 patients with traumatic aortic rupture from vehicular accidents treated between 1971 and 1998 were reviewed. Patients were grouped according to four periods: group 1, 1971 to 1975, (n = 31); group 2, 1976 to 1985, (n = 83); group 3, 1986 to 1994, (n = 82); and group 4, 1994 to 1998 (n = 67). Seventy-one patients died of exsanguination before definitive care. One hundred-ninety two patients had surgical repair with the following techniques: clamp and sew, 6 in group 1, 22 in group 2, 54 in group 3, none in group 4; shunt, 23 in group 1, 39 in group 2, 2 in group 3; cardiopulmonary bypass, 2 in group 1, 1 in group 3. Forty-three patients had partial bypass with the centrifugal pump and heparin-coated circuits in group 4. RESULTS: Operative mortality was 6 of 31 (19%) in group 1, 22 of 61 (36%) in group 2, 15 of 57 (26%) in group 3, and 7 of 43 (16%) in group 4. There was one case of paraplegia in group 1 (4%), ten in group 2 (18%), 11 in group 3 (26%), and none in group 4. This difference of paraplegia between the groups was significant (p<0.002). Significant factors for paraplegia were intraoperative hypotension (p<0.000002), cross-clamp time longer than 30 minutes (p<0.008), pump versus no pump (p<0.008), and younger age group (28+/-11 versus 39+/-17 years) (p<0.03). CONCLUSIONS: There were no statistically significant improvements in mortality rate over the four periods, although, the mortality rate was lowest in the last period when partial bypass with the centrifugal pump was used exclusively. Further, the use of the centrifugal pump with heparin-coated circuits, with femoral vein cannulation into the right atrium and distal aortic perfusion, reduced paraplegia significantly.

Mechanical dysfunction in the border zone of an ovine model of left ventricular aneurysm.

BACKGROUND: The pathophysiology of regional mechanical dysfunction in the border zone (BZ) region of left ventricular aneurysm was studied in an ovine model using magnetic resonance imaging tissue-tagging and regional deformation analysis. METHODS: Transmural infarcts were created in adult Dorsett sheep (n = 8) by ligation of the distal homonymous coronary artery and were allowed to mature into left ventricular aneurysms for 8 to 12 weeks. Animals were imaged subsequently using double oblique magnetic resonance imaging with radiofrequency tissue tagging. Short axis slices were selected for analysis that included predominantly the septal component of the aneurysm as well as adjacent BZ regions in the anterior and posterior ventricular walls. Dark grid patterns of magnetic presaturations were placed on the myocardium and tracked as they deformed during the diastolic, isovolumic systolic, and systolic ejection phases of the cardiac cycle. Regional ventricular wall strains were calculated in BZ regions and regions remote from the aneurysm and compared with strains measured in corresponding regions from normal control sheep (n = 6). RESULTS: Diastolic midwall circumferential strains (fiber extensions) were relatively preserved, but abnormal circumferential lengthening strains were observed in the BZ regions during isovolumic systole. Peak circumferential strains ranged from 0.04 to 0.07 in the BZ regions but averaged -0.05 in the normal hearts (p = 0.002 for the anterior BZ and p = 0.001 for the posterior BZ). Midwall end-systolic fiber strains were depressed in the anterior BZ (-0.03 to -0.09 for the BZ versus -0.11 for the normal heart, p < 0.0001) but not in the posterior BZ (p = 0.19). CONCLUSIONS: Our data support the theory that the stretching of BZ fibers during isovolumic systole contributed to a reduction in fiber shortening during systolic ejection and thus reduced the overall contribution of these fibers to forward ventricular output.

Experience with spiral computed tomography as the sole diagnostic method for traumatic aortic rupture.

BACKGROUND: Spiral computed tomographic (CT) scan is an excellent screen for aortic trauma. Traditionally, aortography is performed when injury is suspected to confirm the diagnosis. We hypothesized that it is safe and expeditious to forgo aortography when the spiral CT demonstrates aortic injury. METHODS: Retrospective review of 54 patients undergoing aortic repair from July 1994 to December 1999. Spiral CT was the initial diagnostic study in 52 patients. Pseudoaneurysm or aortic wall defect in the presence of mediastinal hematoma was considered diagnostic. Angiography, initially routine, was later performed only when requested by the surgeon, and for all "nonnegative" studies (periaortic hematoma without detectable aortic injury). RESULTS: Twenty-six patients underwent angiography before operation (group 1). Nineteen group 1 spiral CTs were unequivocally diagnostic; 7 were nonnegative and angiography was required. Twenty-eight other patients underwent repair based on spiral CT alone (group 2). There was one false-positive result in both groups. There were no unexpected operative findings. Mean time from admission to diagnosis was 5.7+/-3.4 hours for group 1 and 1.7+/-1.7 hours for group 2 (p < 0.01). CONCLUSIONS: Operating on the basis of a diagnostic spiral CT is safe and expeditious. Aortography may be reserved for those with equivocal studies.

The effect of angiotensin II on mitogen-activated protein kinase in human cardiomyocytes.

The role of angiotensin II (Ang II)-receptors on mitogen-activated protein kinase (MAPK) activation in cardiomyocytes remains controversial. Therefore, the current study was designed to investigate the actions of AT(1)- and AT(2)-receptors on Ang II-induced extracellular signal-regulated kinase (ERK), p38 and the c-Jun N-terminal kinase (JNK) MAPK activities in human cardiomyocytes. Human cardiac tissue was obtained from open-heart surgery (n=6). The cardiac tissue was minced and incubated in the special tissue culture system for 24 hours in the absence or presence of Ang II (10(-7) M). These studies were repeated with the AT(1)-receptor antagonist losartan (10(-6) M) or the AT(2)-receptor antagonist PD-123319 (10(-6) M). Immunohistochemical staining and Western blot analysis with phospho-antibodies were performed to determine ERK, JNK and p38 activities. Ang II increased ERK and p38 activities in human cardiomyocytes. The effects of Ang II were abolished by losartan and enhanced by PD-123319. Co-incubation with both losartan and PD-123319 resulted in a decrease of ERK and p38 activities in cardiomyocytes. The immunohistochemical staining of JNK showed no significant differences between Ang II alone, Ang II plus losartan and Ang II plus PD-123319 groups. In conclusion, Ang II has a potent effect on ERK and p38 MAPK activities in cardiomyocytes, by acting through AT(1)-receptors. This effect of Ang II is modified by AT(2)-receptors. Therefore, Ang II, via AT(1)- and AT(2)-receptor stimulation, has a distinct effect on MAPK activity in cardiomyocytes.

Ventricular interaction in the pathologic heart. A model based study.

The effects of direct ventricular interaction and interaction mediated by the pericardium on the diastolic left ventricle (LV) were quantified using idealized models of five pathologic conditions. Two-dimensional (2D) mathematical models were constructed in long and short axis views of four pathologic LV conditions and the normal heart (NL): dilated cardiomyopathy (DCM), concentric LV hypertrophy (HYP), chronic anterior-apical infarction in a normal shaped LV (CAINL), and CAI in a dilated LV (CAID). To assess the effects of RV pressure increase on the LV mechanical state, RV pressure was systematically increased for several LV pressures and changes in the LV diastolic pressure-area relationships, and LV free wall and septal principal stresses and strains were quantified. At higher RV pressures, with pericardial effects included in the models, the pressure-area relationship was similar for all models, indicating that, at these higher pressures, the effects of RV and pericardial pressures are more important than global LV shape, wall thickness, or material properties in determining the pressure-area relationship. There were significant differences among models in the changes in LV free wall and septal stress and strain after an increase in RV pressure. These models may be of use in predicting interaction in the corresponding clinical state.

A multimedia-based histology laboratory course: elimination of the traditional microscope laboratory.

UNLABELLED: We have developed a multimedia-based laboratory course which has enabled us to eliminate the microscope and traditional microscope laboratory that have been mainstays of our histology course and histology courses at almost all institutions where histology is taught. The multimedia laboratory uses a library of histology images (approximately 24,000) stored on videodisc ( HISTOLOGY: A Photographic Atlas, by S. Downing) as its microscope slide collection and accesses those images through barcode and computer interfaces. The laboratory workstations consist of a videodisc player, videodisc monitor, computer, and computer monitor. One workstation is available for every 4-students, and our students are encouraged to work together in groups of four or five. In our current set-up, the students are introduced to and instructed in the basic principles of histology using a computer program that interfaces with the videodisc images. The computer program is divided into 19 chapters (the chapters are typical of the chapters found in a normal histology textbook) and has: (1) a laboratory component that covers the material traditionally covered in the microscope laboratory, and (2) a lecture component that enables the students to evaluate their understanding of the lecture material in a non-punishing way. The laboratory section of each chapter is divided into a "MicroLab" section, an "InFo Time" section, and a "Quiz Time" section. Each of these sections interfaces with histological images stored on the videodisc. The students are encouraged to work through the "MicroLab" section of each chapter before moving on to the "InFo Time" and "Quiz Time" sections. The "MicroLab" sections introduce the students to the various tissues and organs of the body and is interfaced with the videodisc player and the histology images stored on the videodisc. These sections describe the basic histological features of the various tissues and organs and give the students access to multiple examples of what they are studying. The "InFo Time" sections bring up specific images and ask the students to think about the images. Information about the images being observed is available if the students want it and the students can flag those images that they found difficult. The Quiz Time section of the program is also interfaced with the videodisc player and provides access to a large number of histology images stored on the videodisc. The "Quiz Time" sections provide non-punishing review questions that the students can study after she has worked her way through the "MicroLab" and "InFo Time" sections. In addition to the use of a computer program to access the histology images stored on videodisc, we use barcodes that address specific images on the histology videodisc in a variety of ways to augment the students' laboratory and lecture experience. The benefits of using multimedia in place of the traditional microscope and microscope slide collection are numerous and include the speed at which specific histological images can be accessed and reviewed (when compared to finding a structure on a glass slide), a significant reduction in the amount of laboratory time needed by the student to learn the same amount of information, the ease of tutoring on a large monitor screen (when compared to trying to discuss a histological structure with a student through the eyepiece of a microscope), the encouragement of group study (which is difficult to do when a student is working 1-on-with a microscope), and the reduction of the number of faculty necessary to cover a typical histology laboratory session. The use of barcodes that address specific videodisc histology images has greatly changed our examination procedures and has significantly expanded the usefulness of the traditional lecture note handouts given to our students.

Metabolic-morphologic events in the integument of the Pacific hagfish (Eptatretus stoutii).

Light- and electron-microscopic autoradiography were used to obtain a coordinated metabolic-morphologic view of some of the events of cellular differentiation that occur across the epidermis of the Pacific hagfish (Eptatretus stoutii) and which enable this animal to secrete copious amounts of mucus. As judged by epidermal incorporation of [3H]-thymidine in vivo, about 98% of DNA replication is confined to the basal three layers of the total of 6--8 layers of cells. Small mucous cells (SMC), the most numerous of the three major cell types involved in mucigenesis, show in vitro and in vivo radioincorporation profiles of [3H]-L-lysine and [3H]-D-glucosamine which differ markedly from those of [3H]-L-fucose and [3H]-D-galactose. Time-course incorporation profiles (mean silver grains/cell and percentage of cells with at least one cluster of silver grains) of [3H]-L-lysine and [3H]-D-glucosamine not only reflected the metabolic activities of cell renewal and differentiation in basally-located cells but also the high mucigenic activity in cells near the epidermal surface. By contrast, [3H]-L-fucose and [3H]-D-galactose were mainly incorporated by the more mature SMC in juxtanuclear regions near Golgi complexes and newly formed secretory vesicles. The intensity of [3H]-fucose labeling appeared proportional to the intensity of histochemical staining of the apical cytoplasm. The prominent capsule, within SMC in basal and lateral regions, which arises from a tight intermingling of tonofilaments, appears to restrict secretory vesicles to apical regions while the cell progressively differentiates and migrates to the epidermal surface. The other mucigenic cell types, large mucous cells and thread cells, each show distinctive differentiation and radioincorporation patterns.