Cell Maceration Research Articles

DNA content, ploidy level determination and genetic variations of okra (Abelmoschus esculentus L.) genotypes

Okra (Abelmoschus esculentus L.) is a minor small crop in Jordan; it has attracted a lot of attention as a substitute for conventionalvegetables throughout the world. There are conflicting reports about chromosome numbers in this species. To determine the ploidylevel of different okra genotypes, okra root tips were treated with HCl maceration, enzymatic maceration, and Carmine acid squashing.Treating cells with HCl didn’t macerate the cell in a way that enables chromosome count. The enzymatic treatment combinationshowed no significant effect on cell maceration. Carmine’s acetic acid squashing method was able to digest the cells but in a way thatall chromosomes from neighboring cells gathered, making it difficult to count them from each cell. Flow cytometry as an alternativeway to assess okra ploidy, was considered as an option. The genome size of okra ranged from 4.11 pg 2C in genotype 43 to 6.27 pg 2Cin genotype 30.

Chemical properties, crystallinity, and fiber biometry of Jabon (Anthocephalus cadamba) wood for pulp raw material: the effect of age and position

Abstract Jabon (Anthocephalus cadamba) is a promising plant species with straight, visually pleasing trunks that can be harvested at a young age as a resource for pulp and paper production due to the rapid growth rate possessed. Therefore, this study aims to investigate the chemical composition and fiber biometry of 3, 6, and 9-year-old Jabon wood, determining their suitability as a pulp and paper raw material. Samples were collected from both stem and branch sections and analyzed according to the Tappi standard. Fiber cell maceration was conducted to enable the precise measurement of fiber dimensions and derivatives. The results showed high cellulose, holocellulose, and lignin content in Jabon wood, with low to moderate levels of extractives and ash. Additionally, the crystallinity index (CrI) increased with age, particularly from 3 to 9 years old. The branch of the plant, with Grade II fiber derivative quality, had lower CrI compared to the stem, while the inner stem showed a lesser value than the outer part. In conclusion, 3-year-old Jabon wood is a potential raw material for pulp and paper production.

Collagen Content and Collagen Fiber Architecture in the Skin of Shamo Chicken, a Japanese Game Fowl.

Collagen content and collagen fiber architecture in the skin of Shamo chickens were compared between sexes and body parts. Cervical, thoracic, dorsal, femoral, and crural skin samples were collected and their collagen content was analyzed. Collagen fiber specimens were prepared for scanning electron microscopy using the cell maceration method with a NaOH solution. Sex differences in collagen content were only observed in the femoral skin of mature chickens, but not in 10-week-old chicks. The difference in collagen content between body parts was obvious; femoral and crural skin had higher collagen content than those of other parts in both sexes. Scanning electron microscopy indicated that the collagen fiber architecture was quite different between the superficial and deep layers in the dermis, with the former consisting of loosely tangled band-like collagen fibers, and the latter composed of thick and dense layers of collagen bundles in a parallel arrangement. The width of collagen fibers in the superficial layer of the dermis differed between sexes in the dorsal, femoral, and crural skin. From these results, it is likely that the difference in collagen content in the femoral skin is not due to sex hormones but other factors, such as mechanical stimulation in daily activity. Additionally, collagen fiber width in the superficial layer is likely related to the difference in collagen content between sexes and between body parts.

Connective Tissue Framework of the Hepatic Ligaments in the Human Liver

Objectives: To demonstrate the three-dimensional structure of the collagen fiber framework in the human liver ligaments and capsule. Methods: We studied the collagen fiber framework of relatively normal human liver specimens using a cell maceration method and scanning electron microscopy. Results: The collagen fibers of the hepatic falciform ligament subdivided into three types depending on the direction and location. The outer collagen fibers of the hepatic teres ligament formed the longitudinal plate, and the inner fibers had a loop-like structure. The coronary ligament contained two parallel collagen bundles toward the hepatic capsule. The hepatic capsule possesses the outer thicker and inner interlaced layers of the collagen fibers. The hepatic ligaments’ outer layer connected with the hepatic capsule collagen fibers, while the inner layer tended to merge with the hepatic lobular parenchyma’s connective tissue. Conclusions: The hepatic ligaments and liver capsule are layered structures of collagen fibers differing in direction. The hepatic ligaments’ outer layer connected with the liver capsule’s collagen fibers and the inner layer merged with the hepatic lobular parenchyma’s connective tissue.

Changes in Phytochemicals and Determination of Optimum Fermentation Time during Black Tea Manufacturing

Black tea processing consists of four steps, namely withering, CTC, fermentation and drying, while cup quality made tea mostly controlled by fermentation step. This study evaluated biochemical changes at different stages of black tea processing in Bangladesh and determined the optimum fermentation time. Samples were collected from different tea processing stages to measure major phytochemicals and time intervals during fermentation to measure theaflavins and thearubigins ratio. Caffeine content was the least susceptible to processing steps. Biochemical changes started at withering, cell maceration and enzymatic oxidation started at CTC processing, thus the major reduction in the reducing sugar (20.46 to 04.95 ppm), catechin (16.88 to 7.95 ppm) and polyphenol (42.30 to 30.73 ppm) occurred here. The significant changes appeared during fermentation when polyphenol content decreased from 44.66 to 18.23 and catechin from 17.41 to 03.98 ppm due to the breakdown of these compounds to theaflavins (TF) and thearubigins (TR). The TF and TR ratio increased with fermentation time, and the highest of 1:8.4 was found at 50 min, which turned into 1:10 in the final product. The made tea quality parameters were comparable or better at fermentation time of 50 min than the quality of the black tea available in market.

Diversity of extracellular matrix morphology in vertebrate skeletal muscle.

Existing data suggest the extracellular matrix (ECM) of vertebrate skeletal muscle consists of several morphologically distinct layers: an endomysium, perimysium, and epimysium surrounding muscle fibers, fascicles, and whole muscles, respectively. These ECM layers are hypothesized to serve important functional roles within muscle, influencing passive mechanics, providing avenues for force transmission, and influencing dynamic shape changes during contraction. The morphology of the skeletal muscle ECM is well described in mammals and birds; however, ECM morphology in other vertebrate groups including amphibians, fish, and reptiles remains largely unexamined. It remains unclear whether a multilayered ECM is a common feature of vertebrate skeletal muscle, and whether functional roles attributed to the ECM should be considered in mechanical analyses of non-mammalian and non-avian muscle. To explore the prevalence of a multilayered ECM, we used a cell maceration and scanning electron microscopy technique to visualize the organization of ECM collagen in muscle from six vertebrates: bullfrogs (Lithobates catesbeianus), turkeys (Meleagris gallopavo), alligators (Alligator mississippiensis), cane toads (Rhinella marina), laboratory mice (Mus musculus), and carp (Cyprinus carpio). All muscles studied contained a collagen-reinforced ECM with multiple morphologically distinct layers. An endomysium surrounding muscle fibers was apparent in all samples. A perimysium surrounding groups of muscle fibers was apparent in all but carp epaxial muscle; a muscle anatomically, functionally, and phylogenetically distinct from the others studied. An epimysium was apparent in all samples taken at the muscle periphery. These findings show that a multilayered ECM is a common feature of vertebrate muscle and suggest that a functionally relevant ECM should be considered in mechanical models of vertebrate muscle generally. It remains unclear whether cross-species variations in ECM architecture are the result of phylogenetic, anatomical, or functional differences, but understanding the influence of such variation on muscle mechanics may prove a fruitful area for future research.

Infection by a coronatine-producing strain of Pectobacterium cacticidum isolated from sunflower plants in Mexico is characterized by soft rot and chlorosis

A novel strain of Pectobacterium, isolated from infected sunflower plants (Helianthus annuus L.) in Mexico was characterized. Inoculated sunflower plants developed both tissue chlorosis and soft-rot on leaves. The broad host range shown by this pathogen, which included members of the Agavaceae, Asteraceae, Brassicaceae and Solanaceae, was characteristic of the genus Pectobacterium. The metabolic profile and molecular data, in addition to the secretion of plant-cell-wall-degrading enzymes, confirmed its identification as a member of the Pectobacterium genus. A phylogenetic tree constructed on the basis of its 16S rDNA sequence revealed a high identity (98 %) with P. cacticidum. Amplification and restriction-fragment-length-polymorphism analysis of internal transcribed sequences further confirmed its classification as P. cacticidum. Similar to strains of P. atrosepticum, P. carotovorum and Dickeya spp., the new P. cacticidum strain FHLGJ22 has a coronafacic ligase gene and produces coronatine, a virulence factor usually associated with phytopathogenic strains of Pseudomonas syringae. Our results suggest that this strain may utilize a dual infection process involving cell maceration and plant toxins, which is synchronized via an unknown mechanism, might confer an adaptive advantage to colonize different plant hosts.

Cell maceration scanning electron microscopy and computer-derived porosity measurements in assessment of connective tissue microstructure changes in the canine myxomatous mitral valve

Canine myxomatous mitral valve disease is associated with changes in the valve extracellular matrix (ECM). The aim of this study was to examine the use of cell macerated scanning electron microscopy (CMSEM) in evaluating ECM changes in a small sample of valves and to quantify these changes using computer-aided image analysis of sample porosity (a measure of structural disorganisation and collagen loss).The distinct layered structure of the de-cellularised matrix could be seen in the normal valve and there were marked changes in layers and ECM organisation as the disease progressed. Clearly visible and quantifiable, statistically significant changes were found in valve porosity across the entire leaflet thickness and particularly in the valve mid and distal zones. All of these changes are presumed to affect the mechanical function of the valve. In conclusion, CMSEM with computed image analysis can be used to visualise and measure tissue structural changes in a quasi-3-dimensional manner in normal and diseased tissues.

The role of collagen arrangement change during tendon healing demonstrated by scanning electron microscopy

The dry weight of tendon tissue is accounted for mainly by collagen fibers. Accordingly, the tendon-healing process primarily involves repair of collagen fibers. During the remodeling phase of tendon healing, newly proliferating collagen fibers are transformed into a mature repaired tendon. Despite the importance of this phenomenon, the details of fibrous rebuilding have not been reported previously. The aim of this study was to visualize the ultrastructural changes and to obtain a clear understanding of the reorganization of the collagen fibers in the tendon repair site, using rat Achilles tendons. We used scanning electron microscopy (SEM) with cell maceration as the main method of analysis. Pretreatment with cell maceration removed the cellular components successfully. This allowed precise visualization of each collagen fiber and the three-dimensional network of the fibers. This study was the first to apply the cell-maceration/SEM method to observe tendon tissue. Seven days after surgery, new collagen fibers grew extensively in the repair site in a random arrangement. Fourteen days after surgery, the collagen fibers began to form an axial arrangement. Near the tendon stump, this change progressed from the outer layer to the core region. On the other hand, in the middle of the repair site, it progressed from the core to the outer layer. Change in the axial arrangement of collagen fibers contributes to the connection between the repair site and the tendon stump and to the separation of the repair site from the paratenon.

The 3-hydroxy-2-butanone pathway is required for Pectobacterium carotovorum pathogenesis.

Pectobacterium species are necrotrophic bacterial pathogens that cause soft rot diseases in potatoes and several other crops worldwide. Gene expression data identified Pectobacterium carotovorum subsp. carotovorum budB, which encodes the α-acetolactate synthase enzyme in the 2,3-butanediol pathway, as more highly expressed in potato tubers than potato stems. This pathway is of interest because volatiles produced by the 2,3-butanediol pathway have been shown to act as plant growth promoting molecules, insect attractants, and, in other bacterial species, affect virulence and fitness. Disruption of the 2,3-butanediol pathway reduced virulence of P. c. subsp. carotovorum WPP14 on potato tubers and impaired alkalinization of growth medium and potato tubers under anaerobic conditions. Alkalinization of the milieu via this pathway may aid in plant cell maceration since Pectobacterium pectate lyases are most active at alkaline pH.

Dermal neoformation during skin wound healing as demonstrated using scanning electron microscopy.

Little is known about how the structures of connective tissue newly form during the wound-healing process. The authors investigated the repair process of excised skin wounds using alkali water cell maceration scanning electron microscopy. The development of the papillary dermis on the granulation surface proceeded toward the center of the wound, coupled with epidermis migration. No papillary dermis was evident on the granulation surface that had yet to be epithelialized. Finally, a layered structure as observed in normal dermis was reconstructed in the scar tissue. In other words, epidermis, basement membrane, and papillary dermis developed on the granulation surface accompanying epidermal migration, and reticular dermis as the result of maturation of granulation tissue. These findings demonstrate that both the papillary dermis and the reticular dermis develop differently after wounding, and that new papillary dermis and new epidermis develops as if completing "a unit" during the epithelialization process. The authors denote this unit the epithelialization unit.

Morphological study of the regeneration mechanism of acetic acid-injured colon crypts in the rat.

The regeneration mechanism of injured rat colonic mucosa with 1% acetic acid was certified in this study. The injured colons were studied periodically on experimental days 1, 3, 5, 7, 15, and 20 with light and scanning electron microscopy. Specimens were examined in paraffin sections stained with hematoxylin and eosin; crypts were isolated with the HCl digestion method; and three-dimensional stromal collagen tissue was prepared with the NaOH cell maceration method. Damage to the mucosal and submucosal layers peaked between the 1st and 3rd days with edema, regeneration, necrosis, and inflammation. The edema and inflammation subsided, and mucosal atrophy and crypt reduction remained at around 1 week. At 2 weeks the mucosa became thick, and crypts showed many branches in their lower two-thirds; and by 3 weeks the mucosa had recovered to almost normal. The ratio of number of crypts at the base and surface was almost 1.5 on the 15th day and 1.0 on the 20th day, suggesting that each branch progresses upward to create an independent crypt. We believe that the fission mechanism plays an important role in crypt repair after acetic acid injury of the colonic mucosa. As the proliferative zone of the colonic crypt is localized at the crypt base, fission of the crypt starting at the base and progressing up to the surface is the most reasonable and efficient mechanism for repair by increasing the number of crypts.

Matrix metalloproteinases and collagen ultrastructure in moderate myocardial ischemia and reperfusion in vivo

Severe ischemic injury or infarction of myocardium may cause activation of matrix metalloproteinases (MMPs) and damage the interstitial matrix. However, it is unknown whether MMP activation and matrix damage occur after moderate ischemia and reperfusion that result in myocardial stunning without infarction, and if so whether such changes contribute to postischemic myocardial expansion and contractile dysfunction. To address these questions, open-chest anesthetized pigs underwent 90 min of regional ischemia (subendocardial blood flow 0.4 +/- 0.1 ml. g(-1). min(-1)) and 90 min of reperfusion. After ischemia plus reperfusion, histological and ultrastructural examination revealed no myocardial infarction or inflammatory cell infiltration. Myocardial MMP-9 content increased threefold with a fourfold increase in the active form (P < 0.001). Myocardial collagenase content doubled (P < 0.01) but remained in latent form. MMP-2 and tissue inhibitors of metalloproteinases were unaffected. Despite increases in MMPs, collagen ultrastructure (assessed by cell maceration scanning electron microscopy) was unaltered. Intracoronary administration of the MMP inhibitor GM-2487 did not prevent or attenuate myocardial expansion (assessed by regional diastolic dimensions at near-zero left ventricular pressure) or contractile dysfunction. We conclude that although moderate ischemia and reperfusion alter myocardial MMP content and activity, these effects do not result in damage to interstitial collagen, nor do they contribute to myocardial expansion or contractile dysfunction.

Myoarchitecture and connective tissue in hearts with tricuspid atresia

ObjectiveTo compare the atrial and ventricular myoarchitecture in the normal heart and the heart with tricuspid atresia, and to investigate changes in the three dimensional arrangement of collagen fibrils.MethodsBlunt dissection...

Collagen fibrillar network in the optic nerve head of normal monkey eyes and monkey eyes with laser-induced glaucoma – A scanning electron microscopic study

Purpose. To examine the three-dimensional organization of collagen fibrils in the lamina cribrosa of normal monkey eyes and monkey eyes with laser-induced glaucoma. Methods. Intraocular pressure elevation and glaucomatous optic discs were obtained in one eye of three adult monkeys by repeated applications of argon laser to the chamber angle. The monkey eyes were enucleated, and the collagen fibrillar network was investigated by scanning electron microscopy after cell maceration with 10% sodium hydroxide and conductive staining. Results. In normal monkey eyes, round to oval shaped regular laminar pores through which axon bundles exited were observed in the lamina cribrosa. The straight, column-like pores or openings were formed by multilayered laminar plates that aligned vertically in parallel with the optic nerves. The surface of the laminar plates was covered by delicate, loosely arranged collagen fibrils. The inner surface of the pores was smooth, made up of well-packed collagen fibers. In glaucomatous eyes, the laminar pores were clogged by tightened collagen fibrils. The inner surface of the pores was irregular, and the pores were narrowed or distorted. Conclusions. Alterations in the three-dimensional organization of collagen fibrils were demonstrated in the optic nerve head of glaucomatous monkey eyes. The architectural changes may affect the flexibility and resilience required of the lamina cribrosa in supporting optic nerve fibers.

Structural changes in intramuscular connective tissue during the fattening of Japanese black cattle: effect of marbling on beef tenderization.

We investigated changes in structures and mechanical properties of the intramuscular connective tissue during the fattening of Japanese Black steers, using the cell maceration method for scanning electron microscopy. During the early fattening period, from 9 to 20 mo of age, collagen fibrils of the endomysium in longissimus muscle associated more closely with each other, and collagen fibers in the perimysium increased in thickness and their wavy pattern became more regular. These changes were closely related to the increase in mechanical strength of the intramuscular connective tissue and resulted in a toughening of the beef during the period. The shear force value of longissimus muscle decreased after 20 mo of age, concomitantly with the rapid increase in the crude fat content. Scanning electron micrographs of the longissimus muscle dissected from 32-mo-old steers clearly showed that the adipose tissues were formed between muscle fiber bundles, that the honeycomb structure of endomysia was partially broken, and that the perimysium separated into thinner collagen fibers. In semitendinosus muscle, in which the crude fat content was lower (P<.05) than that in longissimus muscle, the structure of the intramuscular connective tissue remained rigid at 32 mo of age. The shear force value of the muscle increased even in the late fattening period, from 20 to 32 mo of age. Thus, the development of adipose tissues in longissimus muscle appears to disorganize the structure of the intramuscular connective tissue and contributes to tenderization of highly marbled beef from Japanese Black cattle during the late fattening period.

Light and scanning electron microscopic study of the palatine mucosa of nine-banded armadillo (Dasypus novemcinctus).

The three-dimensional structure of the lamina propria of the hard and soft palatine mucosa of the nine-banded armadillo was observed by scanning electron microscopy. Sodium hydroxide cell maceration method was applied to demonstrate the architecture of the connective tissue papillae. The palatine mucosa of the armadillo had a triangular shape and measured appr. 6.5 cm length. The hard palate showed 9 transverse palatine plicae while the soft palate was smooth. In the 10% NaOH treated specimens, the lamina propria of the hard palatine mucosa showed numerous connective tissue papillae with a general finger-like shape. These structures were composed by a meshwork of collagen fibers arranged in several directions. On the other hand, the connective tissue papillae of the soft palate mucosa were scattered and small. Numerous openings of glandular ducts with circular or elliptical shape were located in the interplicae area and in the soft palate.

Three-Dimensional Scanning Electron Microscopic Study of Keratoconus Corneas

To examine the 3-dimensional collagen fibril organization in the Bowman layer of keratoconus corneas. Eight keratoconus corneas, 8 corneas with other diseases, and 5 normal human corneas were studied. A cell maceration method in combination with scanning electron microscopy was used to examine the collagen network in the Bowman layer. In normal corneas, the surface of the Bowman layer was smooth and collagen fibrils were regularly arranged. By contrast, sharply edged defects in the Bowman layer were found in keratoconus corneas. Lattice-like configurations of the ruptured Bowman layer and collagenous scar tissue were observed, to varying degrees, in all keratoconus corneas examined. None of the other diseased corneas exhibited the ruptures. Scanning electron microscopy demonstrated alterations in the Bowman layer specific to keratoconus. Fragmentation of the Bowman layer may be an early change leading to keratoconus conditions.

The collagen skeleton of the human umbilical cord at term. A scanning electron microscopy study after 2N-NaOH maceration.

The organization of the collagen fibrils in the human umbilical cord at term is directly visualized by means of a scanning electron microscopy cell maceration method. This technique clearly reveals that there is a much more extensive collagen fibrillar architecture within the umbilical cord than that reported in the classical histological descriptions. The Wharton's jelly, in fact, appears as a spongy network of interlacing collagen fibres and small woven bundles apparently arranged at random and forming a continuous soft skeleton that encases the umbilical vessels. The collagen fibrillar network shows the presence of a wide system of interconnected cavities consisting of canalicular-like structures as well as cavernous and perivascular spaces. This system of cavities might play a mechanical role allowing the storing of the ground substance of the jelly and its diffusion during twisting or compression. Furthermore, it may have an important role facilitating the diffusion throughout the jelly of diffused materials (i.e. water and trophic metabolites) either from or to the umbilical vessels and the amniotic cavity, thus overcoming the lace of a proper vasculature with the jelly.

A morphological study on the histogenesis of human colorectal hyperplastic polyps

Background & Aims Little is known about the histogenesis of human colorectal hyperplastic polyp, although this polyp is clinically very common. Therefore, the structural features of the polyp and their implications regarding histogenesis were studied. Methods A total of 261 foci were examined using scanning electron-microscopic observation of the isolated crypt and surface structure, NaOH cell maceration and scanning electron microscopy, dissecting microscopy, and standard histological analysis. Results In surface view, each polyp crypt was discretely demarcated as in the normal crypt, suggesting that the crypt epithelium had not replaced the adjoining crypt. Notches at the base and various stages of branching, observed in 21.8% of the isolated crypts, were considered to reflect crypt fission. Several polyps with a single crypt mouth consisting of fissioned multiple crypts suggested polyp origin from a single crypt and growth by fission. Juxtaposition of small polyps and their fusion suggested polycentric origin. Almost all polyps showed increased stromal inflammatory cell infiltration and/or a lymphoid follicle at the base. Conclusions Hyperplastic polyps originate by the apparent fusion of single abnormal crypts within a small region of mucosa. The polyps grow by fission of the crypt and fusion of the polycentrically originated polyps. Chronic inflammation has some relation to this process.