, Volume 60, Issue 12, pp 2495–2503 | Cite as

Patterns of cutaneous nerve fibre loss and regeneration in type 2 diabetes with painful and painless polyneuropathy

  • Gidon J. Bönhof
  • Alexander Strom
  • Sonja Püttgen
  • Bernd Ringel
  • Jutta Brüggemann
  • Kálmán Bódis
  • Karsten Müssig
  • Julia Szendroedi
  • Michael Roden
  • Dan Ziegler



The determinants and mechanisms of the development of diabetic sensorimotor polyneuropathy as a painful (DSPN+p) or painless (DSPN-p) entity remain unclear. We examined the degree of cutaneous nerve fibre loss and regeneration in individuals with type 2 diabetes with DSPN+p or DSPN-p compared with individuals with recent-onset type 2 diabetes and corresponding healthy volunteers.


In this cross-sectional study, skin biopsies taken from the distal lateral calf were obtained from individuals with recent-onset type 2 diabetes (n = 32) from the German Diabetes Study, with DSPN+p (n = 34) and DSPN-p (n = 32) from the PROPANE study, and volunteers with normal glucose tolerance (n = 50). Double immunofluorescence staining for protein gene product 9.5 (PGP9.5) (pan-neuronal marker) and growth-associated protein 43 (GAP-43) (nerve regeneration marker) was applied to assess intraepidermal nerve fibre density (IENFD) and length (IENFL) and dermal nerve fibre length (DNFL). DSPN was diagnosed using the modified Toronto Consensus (2011) criteria, while neuropathic pain was assessed using an 11-point Numerical Rating Scale.


After adjustment for age, sex, BMI and HbA1c, IENFD and IENFL were reduced for both markers in individuals with recent-onset diabetes and both DSPN groups compared with control participants (all p < 0.05), but did not differ between the DSPN groups. The DNFL GAP-43/PGP9.5 ratio was higher in the DSPN+p and DSPN-p groups compared with control participants (1.18 ± 0.28 and 1.07 ± 0.10 vs 1.02 ± 0.10; p ≤ 0.05) and in the DSPN + p group compared with DSPN-p (p < 0.05). Correlation analyses showed distinct inverse associations between the DNFL GAP-43/PGP9.5 ratio and PGP9.5 positive IENFD as well as DNFL (IENFD: β = −0.569, DNFL: β = −0.639; both p < 0.0001) in individuals with type 2 diabetes, but not in the control group. A similar pattern was found for correlations between the DNFL GAP-43/PGP9.5 ratio and peripheral nerve function tests.


Dermal nerve fibre regeneration is enhanced in DSPN, particularly in DSPN+p, and increases with advancing intraepidermal nerve fibre loss. These data suggest that, despite progressive epidermal fibre loss, dermal nerve repair is preserved, particularly in DSPN+p, but fails to adequately counteract epidermal neurodegenerative processes.


Nerve regeneration Neuropathic pain Neuropathy Skin biopsy Type 2 diabetes 



Dermal nerve fibre length


Diabetic sensorimotor polyneuropathy


Painful diabetic sensorimotor polyneuropathy


Painless diabetic sensorimotor polyneuropathy


Growth-associated protein-43


German Diabetes Study


Intraepidermal nerve fibre density


Intraepidermal nerve fibre length


Nerve conduction velocity


Neuropathy Disability Score


Numerical Rating Scale


Neuropathy Symptom Score


Protein gene product 9.5


Probing the Role of Sodium Channels in Painful Neuropathies


Small fibre neuropathy


Sensory nerve action potential


Thermal detection threshold


Vibration perception threshold



The authors wish to thank the staff of the Research Group Neuropathy, Institute for Clinical Diabetology at the German Diabetes Center (DDZ), Düsseldorf, Germany, especially F. Battiato, N. Reuß and M. Schroers-Teuber, for their excellent work. The GDS Group consists of A.E. Buyken (Department of Sports and Health, Paderborn University, Paderborn, Germany), G. Geerling (Department of Ophthalmology, Medical Faculty, Heinrich Heine University, Düsseldorf, Germany), J. Eckel, H. Al-Hasani, C. Herder, A. Icks, J. Kotzka, O. Kuss, E. Lammert, D. Markgraf, K. Müssig, W. Rathmann, J. Szendroedi, D. Ziegler and M. Roden (speaker) (all DDZ). Some of the data were presented as an abstract at the 52nd EASD Annual Meeting in Munich in 2016.

Data availability

The data sets generated during and/or analysed during the current study are not publicly available, since they are subject to national data protection laws and restrictions imposed by the ethics committee to ensure data privacy of the study participants. However, they can be applied for through an individual project agreement with PROPANE and/or GDS.


The GDS was initiated and financed by the German Diabetes Center, which is funded by the German Federal Ministry of Health (Berlin, Germany), the Ministry of Innovation, Science, Research and Technology of the state North Rhine-Westphalia (Düsseldorf, Germany), grants from the German Federal Ministry of Education and Research (BMBF) to the German Center for Diabetes Research (DZD), and partly funded through an EFSD award supported by Novartis to DZ and AS. The PROPANE study was initiated by the PROPANE consortium and received funding from the European Union Seventh Framework Programme FP7/2007-2013 (grant no. 602273).

Duality of interest

The authors declare that there is no duality of interest associated with this manuscript.

Contribution statement

All authors were involved in revising the manuscript critically for important intellectual content and gave final approval of the version to be published. GJB contributed to acquisition, analysis and interpretation of data and wrote the manuscript. AS and SP contributed to the acquisition, analysis and interpretation of data, BR and JB contributed to the acquisition of data. KB, KM, JS and MR contributed to the analysis and interpretation of data. DZ contributed to conception and design of the study and to the analysis and interpretation of data. DZ is the guarantor of this work and, as such, had full access to all the data in the study and takes responsibility for the integrity of the data and the accuracy of the data analysis.


  1. 1.
    Happich M, John J, Stamenitis S, Clouth J, Polnau D (2008) The quality of life and economic burden of neuropathy in diabetic patients in Germany in 2002—results from the Diabetic Microvascular Complications (DIMICO) study. Diabetes Res Clin Pract 81:223–230CrossRefPubMedGoogle Scholar
  2. 2.
    Ziegler D, Papanas N, Vinik AI, Shaw JE (2014) Epidemiology of polyneuropathy in diabetes and prediabetes. Handb Clin Neurol 126:3–22CrossRefPubMedGoogle Scholar
  3. 3.
    Landowski LM, Dyck PJ, Engelstad J, Taylor BV (2016) Axonopathy in peripheral neuropathies: mechanisms and therapeutic approaches for regeneration. J Chem Neuroanat 76:19–27CrossRefPubMedGoogle Scholar
  4. 4.
    Yasuda H, Terada M, Maeda K et al (2003) Diabetic neuropathy and nerve regeneration. Prog Neurobiol 69:229–285CrossRefPubMedGoogle Scholar
  5. 5.
    Zochodne DW (2012) The challenges and beauty of peripheral nerve regrowth. J Peripher Nerv Syst 17:1–18CrossRefPubMedGoogle Scholar
  6. 6.
    Spallone V, Greco C (2013) Painful and painless diabetic neuropathy: one disease or two? Curr Diab Rep 13:533–549CrossRefPubMedGoogle Scholar
  7. 7.
    Tesfaye S, Vileikyte L, Rayman G et al (2011) Painful diabetic peripheral neuropathy: consensus recommendations on diagnosis, assessment and management. Diabetes Metab Res Rev 27:629–638CrossRefPubMedGoogle Scholar
  8. 8.
    Lauria G, Lombardi R, Camozzi F, Devigili G (2009) Skin biopsy for the diagnosis of peripheral neuropathy. Histopathology 54:273–285CrossRefPubMedGoogle Scholar
  9. 9.
    Day IN, Thompson RJ (2010) UCHL1 (PGP 9.5): neuronal biomarker and ubiquitin system protein. Prog Neurobiol 90:327–362CrossRefPubMedGoogle Scholar
  10. 10.
    Ziegler D, Papanas N, Zhivov A et al (2014) Early detection of nerve fiber loss by corneal confocal microscopy and skin biopsy in recently diagnosed type 2 diabetes. Diabetes 63:2454–2463CrossRefPubMedGoogle Scholar
  11. 11.
    Kennedy WR, Wendelschafer-Crabb G, Johnson T (1996) Quantitation of epidermal nerves in diabetic neuropathy. Neurology 47:1042–1048CrossRefPubMedGoogle Scholar
  12. 12.
    Denny JB (2006) Molecular mechanisms, biological actions, and neuropharmacology of the growth-associated protein GAP-43. Curr Neuropharmacol 4:293–304CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Benowitz LI, Routtenberg A (1997) GAP-43: an intrinsic determinant of neuronal development and plasticity. Trends Neurosci 20:84–91CrossRefPubMedGoogle Scholar
  14. 14.
    Verze L, Viglietti-Panzica C, Maurizo S, Sica M, Panzica G (2003) Distribution of GAP-43 nerve fibers in the skin of the adult human hand. Anat Rec A Discov Mol Cell Evol Biol 272:467–473CrossRefPubMedGoogle Scholar
  15. 15.
    Fantini F, Johansson O (1992) Expression of growth-associated protein 43 and nerve growth factor receptor in human skin: a comparative immunohistochemical investigation. J Invest Dermatol 99:734–742CrossRefPubMedGoogle Scholar
  16. 16.
    Narayanaswamy H, Facer P, Misra VP et al (2012) A longitudinal study of sensory biomarkers of progression in patients with diabetic peripheral neuropathy using skin biopsies. J Clin Neurosci 19:1490–1496CrossRefPubMedGoogle Scholar
  17. 17.
    Cheng HT, Dauch JR, Porzio MT et al (2013) Increased axonal regeneration and swellings in intraepidermal nerve fibers characterize painful phenotypes of diabetic neuropathy. J Pain 14:941–947CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Bursova S, Dubovy P, Vlckova-Moravcova E et al (2012) Expression of growth-associated protein 43 in the skin nerve fibers of patients with type 2 diabetes mellitus. J Neurol Sci 315:60–63CrossRefPubMedGoogle Scholar
  19. 19.
    Polydefkis M, Hauer P, Sheth S, Sirdofsky M, Griffin JW, McArthur JC (2004) The time course of epidermal nerve fibre regeneration: studies in normal controls and in people with diabetes, with and without neuropathy. Brain 127:1606–1615CrossRefPubMedGoogle Scholar
  20. 20.
    ADA (2012) Diagnosis and classification of diabetes mellitus. Diabetes Care 35(Suppl 1):S64–S71Google Scholar
  21. 21.
    Tesfaye S, Boulton AJ, Dyck PJ et al (2010) Diabetic neuropathies: update on definitions, diagnostic criteria, estimation of severity, and treatments. Diabetes Care 33:2285–2293CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Dworkin RH, Turk DC, Peirce-Sandner S et al (2010) Research design considerations for confirmatory chronic pain clinical trials: IMMPACT recommendations. Pain 149:177–193CrossRefPubMedGoogle Scholar
  23. 23.
    Szendroedi J, Saxena A, Weber KS et al (2016) Cohort profile: the German Diabetes Study (GDS). Cardiovasc Diabetol 15:59CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Lauria G, Ziegler D, Malik R et al (2014) The role of sodium channels in painful diabetic and idiopathic neuropathy. Curr Diab Rep 14:538CrossRefPubMedGoogle Scholar
  25. 25.
    Young MJ, Boulton AJ, MacLeod AF, Williams DR, Sonksen PH (1993) A multicentre study of the prevalence of diabetic peripheral neuropathy in the United Kingdom hospital clinic population. Diabetologia 36:150–154CrossRefPubMedGoogle Scholar
  26. 26.
    Lauria G, Hsieh ST, Johansson O et al (2010) European Federation of Neurological Societies/Peripheral Nerve Society guideline on the use of skin biopsy in the diagnosis of small fiber neuropathy. Report of a joint task force of the European Federation of Neurological Societies and the Peripheral Nerve Society. Eur J Neurol 17(903–912):e944–e909Google Scholar
  27. 27.
    McCarthy BG, Hsieh ST, Stocks A et al (1995) Cutaneous innervation in sensory neuropathies: evaluation by skin biopsy. Neurology 45:1848–1855CrossRefPubMedGoogle Scholar
  28. 28.
    Lauria G, Cazzato D, Porretta-Serapiglia C et al (2011) Morphometry of dermal nerve fibers in human skin. Neurology 77:242–249CrossRefPubMedGoogle Scholar
  29. 29.
    Cheng C, Guo GF, Martinez JA, Singh V, Zochodne DW (2010) Dynamic plasticity of axons within a cutaneous milieu. J Neurosci 30:14735–14744CrossRefPubMedGoogle Scholar
  30. 30.
    Ebenezer GJ, O'Donnell R, Hauer P, Cimino NP, McArthur JC, Polydefkis M (2011) Impaired neurovascular repair in subjects with diabetes following experimental intracutaneous axotomy. Brain 134:1853–1863CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    Costigan M, Scholz J, Woolf CJ (2009) Neuropathic pain: a maladaptive response of the nervous system to damage. Annu Rev Neurosci 32:1–32CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Orstavik K, Namer B, Schmidt R et al (2006) Abnormal function of C-fibers in patients with diabetic neuropathy. J Neurosci 26:11287–11294CrossRefPubMedGoogle Scholar
  33. 33.
    Navarro X, Vivo M, Valero-Cabre A (2007) Neural plasticity after peripheral nerve injury and regeneration. Prog Neurobiol 82:163–201CrossRefPubMedGoogle Scholar
  34. 34.
    Xie W, Strong JA, Zhang JM (2017) Active nerve regeneration with failed target reinnervation drives persistent neuropathic pain. eNeuro.
  35. 35.
    Scheytt S, Riediger N, Braunsdorf S, Sommer C, Uceyler N (2015) Increased gene expression of growth associated protein-43 in skin of patients with early-stage peripheral neuropathies. J Neurol Sci 355:131–137CrossRefPubMedGoogle Scholar
  36. 36.
    Vlckova-Moravcova E, Bednarik J, Dusek L, Toyka KV, Sommer C (2008) Diagnostic validity of epidermal nerve fiber densities in painful sensory neuropathies. Muscle Nerve 37:50–60CrossRefPubMedGoogle Scholar
  37. 37.
    Krishnan ST, Quattrini C, Jeziorska M, Malik RA, Rayman G (2009) Abnormal LDIflare but normal quantitative sensory testing and dermal nerve fiber density in patients with painful diabetic neuropathy. Diabetes Care 32:451–455CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Themistocleous AC, Ramirez JD, Shillo PR et al (2016) The Pain in Neuropathy Study (PiNS): a cross-sectional observational study determining the somatosensory phenotype of painful and painless diabetic neuropathy. Pain 157:1132–1145CrossRefPubMedPubMedCentralGoogle Scholar
  39. 39.
    Sorensen L, Molyneaux L, Yue DK (2006) The relationship among pain, sensory loss, and small nerve fibers in diabetes. Diabetes Care 29:883–887CrossRefPubMedGoogle Scholar
  40. 40.
    Rage M, Van Acker N, Facer P et al (2010) The time course of CO2 laser-evoked responses and of skin nerve fibre markers after topical capsaicin in human volunteers. Clin Neurophysiol 121:1256–1266CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany 2017

Authors and Affiliations

  • Gidon J. Bönhof
    • 1
  • Alexander Strom
    • 1
    • 2
  • Sonja Püttgen
    • 1
  • Bernd Ringel
    • 1
  • Jutta Brüggemann
    • 1
  • Kálmán Bódis
    • 1
  • Karsten Müssig
    • 1
    • 2
    • 3
  • Julia Szendroedi
    • 1
    • 2
    • 3
  • Michael Roden
    • 1
    • 2
    • 3
  • Dan Ziegler
    • 1
    • 2
    • 3
  1. 1.Institute for Clinical Diabetology, German Diabetes Center (DDZ)Leibniz Center for Diabetes Research at Heinrich Heine UniversityDüsseldorfGermany
  2. 2.German Center for Diabetes Research (DZD)MunichGermany
  3. 3.Division of Endocrinology and Diabetology, Medical FacultyHeinrich Heine UniversityDüsseldorfGermany

Personalised recommendations