Lectin Staining of Extensor Digitorum Longus Muscle Cell Membranes in Alloxan Diabetic Rats

Abstract

Aim: Effect of alloxan-diabet on Extensor digitorum longus (EDL) skeletal muscle of rats were observed by lectin staining techniques in light microscope.

Material and Methods: After 30 days alloxan (dose of alloxan: 55 mg/kg) injected by intravenously, samples of muscles were obtained from control and diabetic rats. The muscle samples sections were cutted by cryostat microtome and stained by four biotinylated lectins [Wheat Germ Agglutinin (WGA), Pea Nut Agglutinin (PNA), Concavalin A (ConA), Griffonia simplicifolia I (GS)]. The lectins were fixed by avidin-peroxidase complex. All lectins were bounded to EDL muscle cell membranes of control and diabetic rats.

Results: GS and WGA lectins were strongly (+++) stained extensor digitorum muscle cell membranes of alloxan-diabetic rats. Not only cell membranes but also cytoplasmic miyofibrills of diabetic muscle cells were stained by GS lectin. PNA was moderate (++) stained diabetic muscle cell membranes. Con A was weakly (+) stained diabetic muscle cell membranes with respect to control cell membranes. According to our findings, alloxan-diabetes altered the molecular structure of glycoproteins in cell membranes of EDL skeletal muscles of rats.

Conclusion: We suggested that this study will contribute to diabetes research to show the damaging effects of diabetes on cell membranes.

Keywords

• Extensor digitorum longus muscle cell membrane,Lectin stainning,Diabetes mellitus,Alloxan

Alloxan Diyabetli Sıçanlarda Extensor Digitorum Longus Kas Hücresi Membranlarının Lektin Boyaması

Abstract

Amaç: Alloksan diyabetin sıçanların Extensor Digitorum Longus (EDL) iskelet kasına etkisi lektin boyama tekniği ile ışık mikroskobunda incelendi.

Gereç ve Yöntemler: İntravenöz yoldan alloksan enjekte edildikten 30 gün sonra, kontrol ve diyabetik grubu sıçanlardan kas örnekleri alındı. Kas örneklerinden kryostatlı mikrotom ile kesitleri alındı ve dört biyotinli lektin [Wheat Germ Agglutinin (WGA), Pea Nut Agglutinin (PNA), Concavalin A (ConA), Griffonia simplicifolia (GS)] ile boyandı. Lektinler, avidin-peroksidaz kompleksi ile tespit edildi. Lektinlerin hepsi kontrol ve diyabetik sıçan grubunun EDL iskelet kasına bağlandı.

Bulgular: GS ve WGA lektinleri alloksan diyabetik sıçanların EDL iskelet kaslarını kuvvetli (+++) boyadı. Özellikle GS lektini sadece hücre zarına değil aynı zamanda kas fibrillerini kuvvetli boyadı. PNA lektini orta derecede (++) EDL kaslarını boyadı. Con A ise kontrol grubuna göre diyabetik kasları daha zayıf (+) boyadı. Elde ettiğimiz bulgulara göre alloksan-diyabet sıçanların EDL iskelet kaslarının hücre zarlarındaki glikoproteinlerin moleküler yapısını değiştirmiştir.

Sonuç: Bu çalışma diyabetin hücre zarlarına hasar verici etkisini göstermek açısından diyabetle yapılan araştırmalara katkıda bulunacaktır.

Keywords

• Extensor digitorum longus iskelet kası hücre zarı,Lektin boyama,Diabetes mellitus,Alloksan

References

1. 1. Deveci D, Şanlı Yılmaz F, Güven Y, Acıel M, Ünal A. Değişik zaman süreçlerinde oluşturulan iskemi ve aşırı yüklemenin kas kütlesi ve lifleri üzerine etkileri. S.Ü. Tıp Fak. Derg 2002;18: 149-156.
2. 2. Russell ST, Rajani S, Dhadda RS, Tisdale MJ. Mechanism of induction of muscle protein loss by hyperglycaemia. Exp Cell Res 2009;315:16–25.
3. 3. Paulus SF, Grossıe J. Skeletal muscle in alloxan-diabetes . A comparison of isometric contractions in fast and slow muscles. Diabetes 1983;32:1035-1039.
4. 4. Lithner F, Bergenheim T, Borssén B. Extensor digitorum brevis in diabetic neuropathy: a controlled evaluation in diabetic patients aged 15-50 years. J Intern Med. 1991;230:449-53.
5. 5. Aughsteen AA, Khair AM, Suleiman AA. Quantitative morphometric study of the skeletal muscles of normal and streptozotocin-diabetic rats. 2006;7:382-9.
6. 6. D’Souza DM, Al-Sajee D, Hawke TJ. Diabetic myopathy:impact of diabetes mellitus on skeletal muscle progenitor cells. Front Physiol 2013;4:379.
7. 7. Nonaka K, Une S, Akiyama J. Heat stres attenuates skeletal muscle atrophy of extensor digitorum longus in streptozotocininduced diabetic rats. Acta Physiol hung 2015;102:293-300.
8. 8. Low Wang CC, Hess CN, Hiatt WR, Goldfine AB. Atherosclerotic cardiovascular disease and heart failure in type 2 diabetes-mechanism, management, and clinical considerations. Circulation 2016;133:2459-2502.

9. 9. Zhao ZH, Watschinger B, Brown CD, Beyer MM, Friedman EA. Variations of susceptibility to alloxan induced diabetes in the rabbit. Horm Metab Res 1987; 19:534-7.
10. 10. Weiss RB. Streptozotocin: A review of its pharmacology, efficacy and toxicity. Canc Treat Rep 1982; 66:427–38.
11. 11. Szkudelski T. The mechanism of alloxan and streptozotocin action in β Cells of the rat pancreas. Physiol 2001;50:536–46.
12. 12. Grossıe J. Contractile and electrical characteristic of extensor muscle from alloxan –diabetic rats. An in vitro study. Diabetes 1982;31:194-202.
13. 13. McGuire M and MacDermott M. The influence of streptozotocin diabetes and metformin on erythrocyte volume and on the membrane potential and the contractile characteristics of the extensor digitorum longus and soleus muscles in rats. J Exp Physiol 1999;84: 1051–1058.
14. 14. Cotter MA, Cameron NE, Lean DR, Robertson S. Effect of long-term streptozotocin diabetes on the contractile and histochemical properties of rat muscles. J Exper Physiol 1989;74:65-74.
15. 15. Sharon, N. Lectin. Sci Amer 1977 ; 236:108-119.
16. 16. Lis H, Sharon N. Lectins as molecules and as tools. Ann Rev Biochem 1986;55:35-67.
17. 17. Sharon N, Lis H. Lectins. Springer Netherlands, 2007;pp:XVIII, 454.
18. 18. Vierbuchen M. Lectin receptors. In Current Topics in Pathology. (Ed. Seifert, G.) Springer-Verlag, Berlin 1991; 83:1- 522.
19. 19. Nıchols WS, Nakamura RMAgglutination and agglutination inhibition assays. In Manual of Clinical Laboratory Immunology. (Ed. Rose, N.R., Friedman, H., Fahey, J.L.) Third Ed. American Soc. For Mikrobiol., Washington D.C. 1986; 49- 56.
20. 20. George S, Oh Y, Lindblom S, Vilain S, Rosa AJM, Francis DH, Brözel VS, Kaushik RS. Lectin binding profile of the small intestine of five-week-old pigs in response to the use of chlortetracycline as a growth promotant and under gnotobiotic conditions. J Anim Sci 2007; 85:1640-1650.
21. 21. Pena SDJ, Gordon BB, Karpati G, Carpenter S. Lectin histochemistry of human skeletal muscle. J Histochem Cytochem 1981;29:542-546.
22. 22. Helliwell TR, Gunhan O, Edwards RH. Lectin binding and desmin expression during necrosis, regeneration, and neurogenic atrophy of human skeletal muscle. J Pathol 1989; 159:43-51.
23. 23. Kirkeby S, Moe D, Bøg-Hansen TC. Fucose expression in skeletal muscle: a lectin histochemical study. Histochem J 1993; 25:619-627.
24. 24. Anadolu R, Erdem C, Erdi H, Taşpınar A. Skuamöz hücreli karsinoma tanısında peanut agglutinin. Türk J Dermatopathol 1992;1:46-50.
25. 25. Danguy A, Genten F, Lectin histochemistry on glycoconjugates of the epidermis and dermal glands of Xenopus laevis (Daudin, 1802). Acta Zool. (Stocholm) 1990; 71:17-24.
26. 26. Lever GS, Alroy J, UccI A, Lever WF. Distribution of carbohyrate residues in normal skin . Arch Dermatol Res, 1984; 276:216-223.
27. 27. Bozkurt Ö, Bilgin MD, Severcan F. The effect of diabetes mellitus on rat skeletal extensor digitorum longus musscle tissue: An FTIR study. Spectroscopy 2007; 21:151-160.
28. 28. Reuterwing CO, Hogg E, Holm J. Salivary glands in longterm alloxan-diabetic rats. A quantitative light and electron microscopic study. Acta Pathol. Microbiol. Immunol. Scand. Sect. A 1987 ; 95:131-136.
29. 29. Dunn MJ, Sewry CA, Dubowitz V. Cytochemical studies of lectin binding by diseased human muscle. J Neurol Sci 1982;55:147-159.
30. 30. Capaldi MJ, Dunn MJ, Sewry CA, Dubawitz V. Binding of Ricinus communis 1 lectin to the muscle cell plasma membrane in diseased muscle. J Neurol Sci 1984; 64:315-324
31. 31. Capaldi MJ, Dunn MJ, Sewry CA, Dubowitz V. Lectin binding in human skeletal muscle: a comparison of 15 different lectins. Histochem J 1985; 17:81-92.
32. 32. Yamagami T, Hosaka M, Mori M. Classification of skeletal muscle fibers by comparison of enzyme histochemistry with lectin binding. Cell Mol Biol 1985; 31:241-249.