On the future contents of a small journal of histochemistry

In the last three years, more than 70,000 scientific articles have been published in peer reviewed journals on the application of histochemistry in the biomedical field: most of them did not appear in strictly histochemical journals, but in others dealing with cell and molecular biology, medicine or biotechnology. This proves that histochemistry is still an active and innovative discipline with relevance in basic and applied biological research, but also demonstrates that especially the small histochemical journals should likely reconsider their scopes and strategies to preserve their authorship. A review of the last three years volumes of the European Journal of Histochemistry, taken as an example of a long-time established small journal, confirmed that the published articles were widely heterogeneous in their topics and experimental models, as in this journal''s tradition. This strongly suggests that a journal of histochemistry should keep its role as a forum open to an audience as broad as possible, publishing papers on cell and tissue biology in a wide variety of models. This will improve knowledge of the basic mechanisms of development and differentiation, while helping to increase the number of potential authors since scientists who generally do not use histochemistry in their research will find hints for the applications of histochemical techniques to novel still unexplored subjects.Key words: Basic and applied histochemistry     

Histochemistry and cell biology: the annual review 2010

This review summarizes recent advances in histochemistry and cell biology which complement and extend our knowledge regarding various aspects of protein functions, cell and tissue biology, employing appropriate in vivo model systems in conjunction with established and novel approaches. In this context several non-expected results and discoveries were obtained which paved the way of research into new directions. Once the reader embarks on reading this review, it quickly becomes quite obvious that the studies contribute not only to a better understanding of fundamental biological processes but also provide use-oriented aspects that can be derived therefrom.     

Lelio Orci,the modern master of morphology

Lelio Orci has made seminal contributions to our understanding of pancreatic islet structure and function. He introduced quantitative criteria to structural analysis in the study of endocrine pancreas in a series of works performed in collaboration with Albert Renold, Roger Unger, and Donald Steiner. Orci has moved islet cell morphology from the primitive era of histochemistry and electron microscopy into the modern era of cell biology, applying the most advanced techniques and covering every aspect of normal and pathological structure–function relationships. In collaboration with James Rothman in New York and Randy Schekman in Berkley, Orci discovered that the transport steps from the endoplasmic reticulum to the Golgi complex, and within the Golgi, are mediated by two sets of vesicles coated with protein envelopes different from clathrin.     

The histochemistry and cell biology vade mecum: a review of 2005–2006

The procurement of new knowledge and understanding in the ever expanding discipline of cell biology continues to advance at a breakneck pace. The progress in discerning the physiology of cells and tissues in health and disease has been driven to a large extent by the continued development of new probes and imaging techniques. The recent introduction of semi-conductor quantum dots as stable, specific markers for both fluorescence light microscopy and electron microscopy, as well as a virtual treasure-trove of new fluorescent proteins, has in conjunction with newly introduced spectral imaging systems, opened vistas into the seemingly unlimited possibilities for experimental design. Although it oftentimes proves difficult to predict what the future will hold with respect to advances in disciplines such as cell biology and histochemistry, it is facile to look back on what has already occurred. In this spirit, this review will highlight some advancements made in these areas in the past 2 years.     

Histochemistry through the years, browsing a long-established journal: novelties in traditional subjects

Histochemical journals represent a traditional forum where methodological and technological improvements can be presented and validated in view of their applications to investigate not only cytology and histology in normal and diseased conditions but to test as well hypotheses on more basic issues for life sciences, such as comparative and evolutionary biology. The earliest scientific journals on histochemistry began their publication in the first half of the '50s of the last century, and their readership did not probably change over the years; rather, the authors' interests may have progressively been changing as well as the main topics of their articles. This hypothesis is discussed, based on the subjects of the article published in the first and last ten years in the European Journal of Histochemistry, as an example of old journal which started publication in 1954, being since then the official organ of the Italian Society of Histochemistry. This survey confirmed that histochemistry has provided and still offers unique opportunities for studying the structure, chemical composition and function of cells and tissues in a wide variety of living organisms, especially when the topological distribution of specific molecular components has diagnostic or predictive significance, as it occurs in human and veterinary biology and pathology. Some subjects (e.g. histochemistry applied to muscle cells or to mineralized tissues) have recently become rather popular, whereas a wider application of the histochemical approach may be envisaged for plant cells and tissues.     

Possibilities for the histochemical study of the nuclei of normal brain cells and in pathology

Possibilities of application of some histochemical methods to studying cell nuclei of brain are reviewed considering the following techniques: hybridization histochemistry on the light and electron microscope levels, immunohistochemistry and immunoelectron microscopy, absorbtion and fluorescence histochemistry of nucleic acids, histones, non-histone proteins of chromatin, and of cell nucleus lipids, electron histochemistry. Besides, some physicochemical and molecular-biological methods are considered. Data on human brain research in the norm and upon various brain disorders are particularly provided.     

Thermodynamics,biochemistry and microscopy for the 21st century

The aim of biology is the study of the properties of cells in living intact animals. Many steps of research procedures in biochemistry change the entropy, and therefore, the free energy of the chemical reactions from those in vivo. Sections prepared for histology, histochemistry, electron microscopy and immunocytochemistry, yield limited information about the original cells in life, although their problems have been documented. Approaches to the biological and histological difficulties are suggested.     

Green fluorescent protein (GFP) fusion constructs in gene therapy research

The history of green fluorescent protein (GFP) as a marker is less than 10 years old, but it has already made a major impact on many areas of natural sciences, especially on cell biology and histochemistry. GFP can be detected in living cells without selection or staining and it can be fused to other proteins to yield fluorescent chimeras. The potential of GFP has also been recognised by gene therapy researchers and various GFP-tagged therapeutic proteins have been constructed. These chimeric proteins have been used to determine the expression level, site and time course of the therapeutic gene, or the correlation between gene transfer rate and therapeutic outcome. This review summarises the status of the applications of GFP fusions in gene therapy research.     

Advantages of histochemistry for the study of cell biology

Summary Bridging between the two bodies of cell science, histochemistry has brought together knowledge of morphological structures and biochemical constituents and provided insight into the function of one or the other. Where it has localized an enzyme, hormone or other entity of known biological activity to a cell type, histochemistry has contributed insight into the cell's function. By detecting heterogeneity in the content of an enzyme or glycoconjugate within a presumed uniform population of cells, the histochemical approach has differentiated among these cells subtypes with demonstrated or presumed differences in function. On the other hand, in instances where it has located an isozyme, antigen, glycoconjugate or other entity of uncertain significance in a cell or organelle of known function, histochemistry has suggested a possible role for the constituent related to that of the structure. Histochemical examination provides the observer with a different view of body tissues and their composition from that obtained by strictly morphological or chemical techniques. Information acquired through this advantage is cited wherein an immunohistochemical method disclosed previously undiscovered neural organs and carbohydrate histochemistry detected previously unrecognized glycoconjugates. Presented as the David Glick Lecture at the 9th International Congress of Histochemistry and Cytochemistry in Maastricht The Netherlands, 30 August–5 September, 1992.     

Hybridization histochemistry

In this review we have used our own recent work as a flagship to illustrate the recent renaissance of interest in hybridization histochemistry. A trickle of papers followed the initial key excursion into the in situ labeling of tissue sections (48-50). Our own entry into this field started in 1978 and since then a confluence of important questions and technical advances has served to make hybridization histochemistry much more attractive as a research tool. Hybridization histochemistry is able to solve some problems for which there is no other suitable technique at this time. Hybridization histochemistry provides the location of anatomical sites of gene expression, and viral replication, with uniquely high specificity. We have taken 32P-labeled probes to what appears to be their limit of resolution, which is single cells in thin sections. While 32P has clear disadvantages, exposure time is relatively short and the use of fast-X-ray film to preview the results and estimate exposure time for emulsion has been turned to advantage. Our introduction (27) of the use of whole-mouse sections in hybridization histochemistry has great potential in hormonal, enzymatic, and growth factor gene expression and will no doubt prove of great use in developmental studies and examination of viral infection. The use of synthetic DNA (synthetic oligonucleotides) unshackles the technique from the need for an associated molecular biology laboratory and at once widens the horizon of application of the technique. Although hybridization histochemistry is a valuable research tool which will soon find a niche in many fields, in a short time it should become a key diagnostic aid. It may well become the method of preference for detection of the expression of oncogenes and other cancer-related genes and for viruses which for other reasons are difficult to detect.     

From chemical neuroanatomy to an understanding of the olfactory system

The olfactory system is the appropriate model for studying several aspects of neuronal physiology spanning from the developmental stage to neural network remodelling in the adult brain. Both the morphological and physiological understanding of this system were strongly supported by classical histochemistry. It is emblematic the case of the Olfactory Marker Protein (OMP) staining, the first, powerful marker for fully differentiated olfactory receptor neurons and a key tool to investigate the dynamic relations between peripheral sensory epithelia and central relay regions given its presence within olfactory fibers reaching the olfactory bulb (OB). Similarly, the use of thymidine analogues was able to show neurogenesis in an adult mammalian brain far before modern virus labelling and lipophilic tracers based methods. Nowadays, a wealth of new histochemical techniques combining cell and molecular biology approaches is available, giving stance to move from the analysis of the chemically identified circuitries to functional research. The study of adult neurogenesis is indeed one of the best explanatory examples of this statement. After defining the cell types involved and the basic physiology of this phenomenon in the OB plasticity, we can now analyze the role of neurogenesis in well testable behaviours related to socio-chemical communication in rodents.     

Histochemical methods for detecting nitric oxide synthase

Summary The three isoforms of nitric oxide synthase (NOS), neuronal (nNOS), endothelial (eNOS), and inducible (iNOS), can be visualized in cells and tissues by NADPH-diaphorase (NADPH-d) histochemistry, immunocytochemistry and in situ hybridization. Histochemical demonstration of NADPH-d shows the formazan final reaction product as a solid blue deposit. The ultrastructural localization of NADPH-d in the rat hippocampus showed an electron-dense deposit on membranes predominantly of the endoplasmic reticulum. The immunohistochemical demonstration of nNOS, using the nickel enhancement technique, shows positive reaction product over the dendrites and the soma of the nerve cell in the rat brain. Ultrastructural localization of nNOS in whole mount preparations of myenteric plexus and circular smooth muscle from guinea-pig ileum shows that NOS immunoreactivity was patchily distributed in myenteric neurones and was not specifically associated with any intracellular organelles or with plasma membranes. In situ hybridization, using radio-labelled probes, was used to study nNOS mRNA in lumbar dorsal root ganglia after peripheral transection of the sciatic nerve in rats. Labelling of the NOS mRNA-positive neurones is observed as a series of dense granules over the entire cell. NADPH-d histochemistry, immunocytochemistry and in situ hybridization each have a significant role to play in the localization of NOS. NADPH-d detects an enzyme associated with the NOS molecule, immunocytochemistry detects the NOS molecule, and in situ hybridization detects mRNA for NOS. Therefore, if each of these techniques is applied in carefully controlled experiments, consideration of the accumulated data should be valuable in revealing insights into the biology of NOS.     

Localization of Acid and Alkaline Phosphatases in Staphylococcus aureus

The localization of acid and alkaline phosphatases in Staphylococcus aureus was studied by fractionation of cells after treatment with the L-11 enzyme and by electron microscopic histochemistry. The two enzyme activities were located in distinctly different positions at the surface of the cells. Acid phosphatase appeared to be localized around the cell membrane of the bacteria, because the enzyme was recovered exclusively in the membrane fraction and because deposition of lead phosphate was detected by electron microscopic histochemistry on the inner surface of the cell membrane of intact bacteria and spheroplasts. The highest specific activity of alkaline phosphatase was also associated with the membrane fraction. However, on electron microscopic histochemistry of intact cells, the deposition of lead phosphate was only seen on the outer surface of the cell wall.     

Identification of peroxidase-positive astrocytes by combined histochemical and immunolabeling techniques in situ and in cell culture.

A subpopulation of astrocytes in the vertebrate brain and in cysteamine-treated brain cell cultures contain cytoplasmic granules which exhibit an affinity for Gomori stains, orange-red autofluorescence, and non-enzymatic endogenous peroxidase activity. Visualization of these cells at the light microscopic level is confounded by the nonspecificity of the various histochemical methods routinely employed. In an attempt to circumvent this problem, we assayed for peroxidase-positive astrocytes using various combinations of diaminobenzidine (DAB) histochemistry and immunolabeling for the astrocyte-specific marker glial fibrillary acidic protein (GFAP). We determined that (a) DAB histochemistry in conjunction with avidin-biotin-immunoperoxidase labeling for GFAP specifically detects peroxidase-positive astrocytes in situ and (b) DAB histochemistry combined with indirect immunofluorescence for GFAP effectively demonstrates these cells in cysteamine-treated brain cell cultures.     

Histochemical Method for Detecting Nitric Oxide Synthase Activity in Cell Cultures

NADPH diaphorase histochemistry has been used extensively for detecting nitric oxide synthase (NOS) activity in various cell types including neuronal cell bodies, vascular endothelium, cells of the immune system and epithelial cells. The use of the diaphorase technique in cell cultures to study the induction of NOS has not been investigated. In this paper we report the use of diaphorase histochemistry as a good marker for the detection of NOS activity in cultured cells. This technique can be used in conjunction with other established techniques to determine the presence and activity of NOS in cultured cells.     

An air-liquid interface promotes the differentiation of gastric surface mucous cells (GSM06) in culture

The gastric surface epithelium is situated at an air-liquid interface because the luminal surface of the alimentary tract is in continuity with the air phase. However, the effects of this microenvironment on the gastric epithelium remain unclear. The aim of this study was to clarify the effects of an air-liquid interface on gastric epithelial cell biology. Gastric surface mucous cells (GSM06) were cultured at an air-liquid interface. Cultured cells were examined by histology, histochemistry, and transmission electron microscopy. When the cells were cultured at an air-liquid interface, the surface cells on the collagen gel became tall columnar and secreted periodic acid-Shiff-positive substances at the apical surface. These cells indicated many mucous granules in the apical cytoplasm and organized the basal lamina at the contact side with the gel. In contrast, under immersed condition, the surface cells showed immature features. This is the first report of an air-liquid interface promoting the differentiation of gastric surface mucous cells in a reconstruction culture of the gastric surface epithelial layer, suggesting that an air-liquid interface may function as a crucial luminal factor to maintain the homeostasis of gastric mucosa.