Immunohistochemical substantiation of the possibility of use electrochemical method whith using nanotechnological biosensors for assessing alkaline phosphatase activity in tissue colorectal carcinoma

Cover Page

Cite item

Full Text

Open Access Open Access
Restricted Access Access granted
Restricted Access Subscription or Fee Access

Abstract

Introduction. Alkaline phosphatase (ALP) is an enzyme from the class of hydrolases, widely present in human tissues and organs. Intestinal ALP is one of the enzyme isoforms that is expressed in the mucous membrane throughout the intestine and is a marker of intestinal epithelial differentiation. It is known that the products of the chemical reaction between intestinal alkaline phosphate and a specific substrate, 1-naphthyl phosphate, have electrochemical activity. This makes it possible to evaluate the activity of the enzyme in biological tissues using the electrochemical method using nanotechnological biosensors.

The aim of the study. To evaluate the diagnostic significance of the electrochemical method for assessing alkaline phosphatase activity by comparing it with the results of histological and immunohistochemical studies in colorectal carcinoma.

Material and methods. A parallel electrochemical and morphological (histological and immunohistochemical with antibodies to intestinal alkaline phosphatase) study of material from colorectal carcinoma and the colon mucosa outside the tumor of 78 patients was carried out.

Results. In 70 patients, the current obtained from electrochemical study of tumor biopsies was significantly lower (49.2 nA (95% CI 41.3–88.9) than in biopsies of the intestinal mucosa outside the tumor (119.7 nA (95% CI 96.8–167.1), p<0.05). Histologically, the tumor tissue was represented by adenocarcinoma of varying degrees of differentiation. An immunohistochemical study revealed that the expression of intestinal ALP was absent in carcinoma cells, while in the epithelium of the colon mucosa outside the neoplasm, pronounced diffuse membrane expression of the enzyme was noted. In 8 patients, there was no association between the results of electrochemical and morphological studies due to the presence of non-tumor tissues in the material. An immunohistochemical study revealed that intestinal alkaline phosphatase can be expressed in immune cells and neurons of the submucosal nerve plexuses.

Conclusion. A comparison of the results of electrochemical, histological and immunohistochemical studies indicates that the electrochemical method has a high diagnostic value and can be used in screening for colorectal carcinoma.

Full Text

Restricted Access

About the authors

Anton N. Belkin

Perm State Medical University named after Academician E.A. Wagner Ministry of Health of the Russian Federation

Author for correspondence.
Email: belkinanton87@gmail.com
ORCID iD: 0000-0002-8646-0483

senior lecturer at the Department of Pathological Anatomy with an autopsy course

Russian Federation, Petropavlovskaya st., 26, Perm, 614000

Henrietta G. Freynd

Perm State Medical University named after Academician E.A. Wagner Ministry of Health of the Russian Federation

Email: gfreynd@mail.ru
ORCID iD: 0000-0002-2861-4878

Head of the Department of Pathological Anatomy with an autopsy course, Doctor of Medical Sciences, Professor

Russian Federation, Petropavlovskaya st., 26, Perm, 614000

Anatoly G. Kochetov

Institute of Laboratory Medicine, Leningradsky Prospekt

Email: kochetov.lab@yandex.ru
ORCID iD: 0000-0003-3632-291X

rector of the Institute, Professor

Russian Federation, 80G, office 911A, Moscow, 117042

References

  1. Rawla P., Sunkara T., Barsouk A. Epidemiology of colorectal cancer: incidence, mortality, survival, and risk factors. Prz Gastroenterol. 2019; 14 (2): 89–103. doi: 10.5114/pg.2018.81072
  2. National Cancer Institute. Surveillance, epidemiology, and end results program (SEER). http://seer.cancer.gov
  3. Muginova S.V., Zhavoronkova A.M., Polyakov A.E., Shekhovtsova T.N. Application of alkaline phosphatases from different sources in pharmaceutical and clinical analysis for the determination of their cofactors; zinc and magnesium ions. Anal Sci. 2007; 23: 357–63. doi: 10.2116/analsci.23.357
  4. Metwalli O.M., Mourand F.E. Studies on organ-specific alkaline phosphatases in relation to their diagnostic value. Z Ernahrung Swiss. 1980; 19: 154–8. doi: 10.1007/BF02018779
  5. Molnár К., Vannay А., Szebeni В., Fanni Bánki N., Sziksz E., Cseh Á., Győrffy H., László Lakatos P., Papp M., Arató A., Veres G. Intestinal alkaline phosphatase in the colonic mucosa of children with inflammatory bowel disease World J. Gastroenterol. 2012; 18: 3254–9. doi: 10.3748/wjg.v18.i25.3254
  6. Narisawa S., Huang L., Iwasaki A., Hasegawa H., Alpers D.H., Millán J.L. Accelerated fat absorption in intestinal alkaline phosphatase knockout mice. Mol. Cell. Biol. 2003; 23: 7525–30. doi: 10.1128/MCB.23.21.7525-7530.2003
  7. Tuin A., Poelstra K., de Jager-Krikken A., Bok L., Raaben W., Velders M. P., Dijkstra G. Role of alkaline phosphatase in colitis in man and rats. Gut. 2009; 58 (3): 379–87. doi: 10.1136/gut.2007.128868
  8. Akiba Y., Mizumori M., Guth P.H., Engel E., Kaunitz J.D. Duodenal brush border intestinal alkaline phosphatase activity affects bicarbonate secretion in rats. Am J Physiol Gastrointest Liver Physiol. 2007; 293 (6): 1223–33. doi: 10.1152/ajpgi.00313.2007
  9. Estaki M., DeCoffe D., Gibson D.L. Interplay between intestinal alkaline phosphatase, diet, gut microbes and immunity. World J. Gastroenterol. 2014; 20 (42): 15650–6. doi: 10.3748/wjg.v20.i42.15650
  10. Chen K.T., Malo M.S., Moss A.K., Zeller S., Johnson P., Ebrahimi F., Mostafa G., Alam S.N., Ramasamy S., Warren H.S., Hohmann E.L., Hodin R.A. Identification of specific targets for the gut mucosal defense factor intestinal alkaline phosphatase. Am. J. Physiol Gastrointest Liver Physiol. 2010; 299 (2): 467–75. doi: 10.1152/ajpgi.00364.2009
  11. Moss D.W. Alkaline phosphatase isoenzymes. Clin Chem. 1982; 28 (10): 2007–16.
  12. Wilkers J.M., Gamer A., Peters T.J. Studies on the localization and properties of rat duodenal HCO3-ATPase with special relation to alkaline phosphatase. Biochim Biophys Acta. 1987; 924 (1): 159–66. doi: 10.1016/0304-4165(87)90083-3
  13. Singh S.B., Lin H.C. Role of Intestinal alkaline phosphatase in innate immunity. Biomolecules. 2021; 11 (12): 1784. doi: 10.3390/biom11121784
  14. Hinnebusch B.F., Siddique A., Henderson J.W., Malo M.S., Zhang W., Athaide C.P., Abedrapo M.A., Chen X., Yang W.V., Hodin R.A. Enterocyte differentiation marker intestinal alkaline phosphatase is a target gene of the gut-enriched Kruppel-like factor. Am. J. Physiol Gastrointest Liver Physiol. 2004; 286 (1): 23–30. doi: 10.1152/ajpgi.00203.2003
  15. Barnard J.A., Warwick G. Butyrate rapidly induces growth inhibition and differentiation in HT-29 cells. Cell Growth Differ. 1993; 4 (6): 495–501.
  16. Kabat E. A., Furth J. A histochemical study of distribution of alkaline phosphatase in various normal and neoplastic tissues. Am. J. Pathol. 1941; 17 (3): 303–18.
  17. Li M., Jiang F., Xue L., Peng C., Shi Z., Zhang Z., Li J., Pan Y., Wang X., Feng C., Qiao D., Chen Z., Luo Q., Chen X. Recent Progress in Biosensors for Detection of Tumor Biomarkers. Molecules. 2022; 27 (21): 7327. doi: 10.3390/molecules27217327
  18. Верник С., Белкин А.Н., Фрейнд Г.Г., Кацнельсон М.Д., Шахам-Диаманд Й., Лицын С.Н., Четвертных В.А. Морфологическое обоснование возможности использования электрохимических биосенсоров в диагностике колоректального рака. Пермский медицинский журнал. 2012; 5: 5–7. [Vernik S., Belkin A.N., Freynd G.G., Katsnelson M.D., Shaham-Diamand Y., Litsyn S.N., Chetvertnykh V.A. Morphological substantiation of the possibility of using electrochemical biosensors in the diagnosis of colorectal cancer. Perm Medical Journal. 2012; 5: 5–7 (in Russian)].
  19. Шелудько В.С. Девяткова Г.И. Теоретические основы медицинской статистики (статистические методы обработки и анализа материалов научно-исследовательских работ): методические рекомендации, 3-е изд., исправленное и дополненное. Пермь: ФГБОУ ВО «ПГМУ имени академика Е.А. Вагнера» Минздрава России; 2016; 80. [Sheludko V.S. Devyatkova G.I. Theoretical foundations of medical statistics (statistical methods of processing and analysis of research materials): methodological recommendations, 3rd ed., corrected and expanded. Perm: Federal State Budgetary Educational Institution of Higher Education “Perm State Medical University named after Academician E.A. Wagner» of the Russian Ministry of Health; 2016; 80 (in Russian)].
  20. Ланг Т.А., Сесик М. Описание статистики в медицине. Руководство для авторов, редакторов и рецензентов. М.: Практическая медицина, 2011; 480. [Lang T.A., Sesik M. Description of statistics in medicine. A guide for authors, editors and reviewers. M.: Practical Medicine, 2011; 480 (in Russian)].
  21. Song Z.M., Brookes S.J., Costa M. Characterization of alkaline phosphatase-reactive neurons in the guinea-pig small intestine. Neuroscience. 1994; 63 (40): 1153–67. doi: 10.1016/0306-4522(94)90580-0
  22. Stewart C. Leukocyte alkaline phosphatase in myeloid maturation. Pathol. 1974; 6 (3): 287–93. doi: 10.3109/00313027409068999
  23. Kumar V., Abbas A., Aster J. Robbins basic pathology 9th edition. Saunders, 2012; 928.

Supplementary files

Supplementary Files
Action
1. JATS XML
Download
2. Fig. 1. Hardware complex for electrochemical research. 1 – mixing device, 2 – set of eight biosensors, in the wells of which pipettes for air supply are immersed, 3 – portable potentiostat

Download (2MB)
Indexing metadata
3. Fig. 2. Histological structure of adenocarcinoma G3 biopsy. a – tumor tissue consists from tubular and focal solid structures of atypical glandular epithelium (H&E stain, ×400); b – immunohistochemical study with monoclonal antibodies to alkaline phosphatase: absence of alkaline phosphatase expression in tumor cells (×400)

Download (1MB)
Indexing metadata
4. Fig. 3. Colon mucosa outside the tumor. а – H&E stain, ×100; b – immunohistochemical study with monoclonal rabbit antibodies to ALP – intensive diffuse expression of ALP in the cytoplasmic membranes of colonocytes (×100)

Download (660KB)
Indexing metadata
5. Fig. 4. Current during electrochemical study of biopsy samples of the colon mucosa (a: 16.82±2.41 nA) and tumor (b: 25.9±16.12 nA)

Download (87KB)
Indexing metadata
6. Fig. 5. Immunohistochemical study with monoclonal rabbit antibodies to ALP. а – moderate diffuse membrane and cytoplasmic expression of alkaline phosphatase in macrophages, lymphocytes, plasma cells, absence of alkaline phosphatase expression in the endothelium of blood vessels (×400); b – moderate focal cytoplasmic expression of alkaline phosphatase in ganglion cells of the submucosal nerve plexus (×400)

Download (2MB)
Indexing metadata

Copyright (c) 2024 Russkiy Vrach Publishing House

This website uses cookies

You consent to our cookies if you continue to use our website.

About Cookies