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Journal of Zhejiang University-SCIENCE B

, Volume 16, Issue 11, pp 948–956 | Cite as

Impact of type 2 diabetes on the plasma levels of vascular endothelial growth factor and its soluble receptors type 1 and type 2 in patients with peripheral arterial disease

  • Radosław WieczórEmail author
  • Grażyna Gadomska
  • Barbara Ruszkowska-Ciastek
  • Katarzyna Stankowska
  • Jacek Budzyński
  • Jacek Fabisiak
  • Karol Suppan
  • Grzegorz Pulkowski
  • Danuta Rość
Article

Abstract

Objective

Type 2 diabetes coexistent with lower extremity artery disease (peripheral arterial disease (PAD)) can be observed in numerous patients. The mechanism compensating for ischemia and contributing to healing is angiogenesis—the process of forming new blood vessels. The purpose of this study was to assess the likely impact of type 2 diabetes on the plasma levels of proangiogenic factor (vascular endothelial growth factor A (VEGF-A)) and angiogenesis inhibitors (soluble VEGF receptors type 1 and type 2 (sVEGFR-1 and sVEGFR-2)) in patients with PAD.

Method

Among 46 patients with PAD under pharmacological therapy (non-invasive), we identified, based on medical history, a subgroup with coexistent type 2 diabetes (PAD-DM2+, n=15) and without diabetes (PAD-DM2−, n=31). The control group consisted of 30 healthy subjects. Plasma levels of VEGF-A, sVEGFR-1, and sVEGFR-2 were measured using the enzyme-linked immunosorbent assay (ELISA) method.

Results

The subgroups of PAD-DM2+ and PAD-DM2−revealed significantly higher concentrations of VEGF-A (P=0.000 007 and P=0.000 000 1, respectively) and significantly lower sVEGFR-2 levels (P=0.02 and P=0.000 01, respectively), when compared with the control group. Patients with PAD and coexistent diabetes tended to have a lower level of VEGF-A and higher levels of sVEGFR-1 and sVEGFR-2 comparable with non-diabetic patients.

Conclusions

The coexistence of type 2 diabetes and PAD is demonstrated by a tendency to a lower plasma level of proangiogenic factor (VEGF-A) and higher levels of angiogenesis inhibitors (sVEGFR-1 and sVEGFR-2) at the same time. Regardless of the coexistence of type 2 diabetes, hypoxia appears to be a crucial factor stimulating the processes of angiogenesis in PAD patients comparable with healthy individuals, whereas hyperglycemia may have a negative impact on angiogenesis in lower limbs.

Keywords

Angiogenesis Peripheral arterial disease Soluble receptors Type 2 diabetes mellitus Vascular endothelial growth factor 

2 型糖尿病对外周动脉疾病患者血浆内的血管内皮生长因子及其可溶性受体浓度的影响

中文概要

目 的

研究2 型糖尿病对外周动脉疾病患者血浆内的血管内皮生长因子(VEGF-A)及其水溶性受体(sVEGFR-1 和sVEGFR-2)浓度的影响。

创新点

首次研究了2 型糖尿病对外周动脉疾病患者血浆内sVEGFR-1 和sVEGFR-2 浓度的影响。

方 法

选取46 个外周动脉疾病患者, 根据有无2 型糖尿病分为糖尿病组(15 例)和无糖尿病组(31 例), 另选30 个健康志愿者为正常对照组。采用酶联免疫吸附法(ELISA)检测他们血浆中VEGF-A及sVEGFR-1 和sVEGFR-2 的浓度, 然后通过对比各组浓度研究2 型糖尿病的影响。

结 论

与正常对照组相比, 外周动脉疾病患者具有较高的VEGF-A 浓度(2 型糖尿病组 P=0.000 007, 非糖尿病组 P=0.000 000 1)以及较低的sVEGFR-2浓度(2 型糖尿病组 P=0.02, 非糖尿病组 P=0.000 01)。同时, 2 型糖尿病组比非糖尿病组具有较低的VEGF-A 浓度及较高的sVEGFR-1 和sVEGFR-2 浓度。研究结果表明: 无论2 型糖尿病是否共存, 缺氧是导致血管生成的一个关键的刺激因素; 同时, 高血糖状态对下肢的血管生成有抑制作用。

关键词

血管生成 外周动脉疾病 可溶性受体 2 型糖尿病 血管内皮生长因子 

CLC number

R587.1 

1 Introduction

Type 2 diabetes and atherosclerosis can be regarded as epidemic diseases at the turn of the 20th and 21st centuries. Coexisting complications of diabetes and lower extremity artery disease (peripheral arterial disease (PAD)) account for the most common causes of lower limb amputation, whereas the frequency of amputations in patients with diabetes is five- to ten-fold higher than that in non-diabetic individuals. Macroangiopathic and microangiopathic complications are a major cause of mortality in diabetes. Symptoms like intermittent claudication occur twice as often in individuals with diabetes, which increases the risk of PAD three to four times (Norgren et al., 2007; Jude et al., 2010).

Vascular endothelial growth factor A (VEGF-A) is considered to be the main proangiogenic factor as it participates in the formation of new blood vessels. In patients with PAD, an angiogenic impulse is ischemia of tissues caused by narrowed vessels as a result of atherosclerotic plaque. Through hypoxia inducible factors (e.g. hypoxia inducible factor 1α (HIF-1α)), endothelial cells produce VEGF-A which participates in several angiogenesis stages. For example, VEGF-A leads to the activation and migration of endothelial cells and inhibits their apoptosis. Additionally, VEGF-A mobilizes endothelial progenitor cells (EPCs) which migrate to ischemic tissues, where they differentiate to endothelial cells and proliferate rapidly (Bao et al., 2009). As clinical and laboratory data indicate, impaired angiogenesis can be observed in, for example, patients with diabetic foot syndrome (DFS) (Drela et al., 2014). Therefore, it is necessary to investigate factors that have an influence on angiogenesis in PAD and diabetes alike. The practical use of discovered mechanisms is emphasized in effective attempts to apply VEGF-A and bone marrow mononuclear cells, for example, in the treatment of critical limb ischemia coexistent with diabetes (Kusumanto et al., 2006; Skóra et al., 2013).

Available studies suggest that soluble vascular endothelial growth factor receptors type 1 and type 2 (sVEGFR-1 and sVEGFR-2) may be regarded as angiogenic inhibitors. The studies by Liu et al. (2014) indicated that sVEGFR-2, when forming complexes with VEGF-A, decreased bioavailability of VEGF-A to endothelial receptors, thus affecting a decreased biological activity of VEGF-A.

In blood of patients with atherosclerosis and diabetic individuals, higher levels of VEGF-A caused by tissue hypoxia can be found (Botti et al., 2012). However, the role of different concentrations of sVEGFR-1 and sVEGFR-2 in the process of angiogenesis is still unknown. The aim of this study was to assess the likely impact of type 2 diabetes on the levels of VEGF-A, sVEGFR-1, and sVEGFR-2 in plasma of patients with PAD.

2 Materials and methods

Among 46 patients (28 males and 18 females), treated in the Clinic of Vascular and Internal Medicine of Dr. Jan Biziel University Hospital No. 2 in Bydgoszcz (Poland) for symptomatic PAD, we identified, based on medical history, a subgroup of 15 patients (8 males and 7 females) with PAD and coexisting diabetes (PAD-DM2+) and a subgroup of 31 non-diabetic individuals (20 males and 11 females) with PAD (PAD-DM2−). Exclusion criteria for participation in the study were a history of neoplastic disease, diabetic retinopathy, and a lack of a patient consent. PAD severity was determined based on the Fontaine classification. The control group consisted of 30 healthy volunteers (15 males and 15 females, average age (55.9±7.7) years), without clinically manifested atherosclerosis, neoplastic disease, or abnormal carbohydrate metabolism.

Venous blood plasma samples were collected to determine concentrations of VEGF-A and its soluble receptors sVEGFR-1 and sVEGFR-2. The enzyme-linked immunosorbent assay (ELISA) method was applied, using Quantikine reagents (R & D Systems, USA). Approval of the study was obtained from the local Bioethics Commission of Ludwik Rydygier Collegium Medicum in Bydgoszcz, Nicolaus Copernicus University in Toruń (CM UMK; Document No. 509/2011), and the clinical investigations were carried out in accordance with the Helsinki Declaration. The consent for participation in the study was freely given, informed, and expressed in writing on a relevant form. The results were statistically analyzed using Statistica Ver. 10.0 software (StatSoft, USA) and Excel (Windows, USA). The significance level was set at P =0.05.

3 Results

Table 1 demonstrates the characteristics of the study group (PAD, n =46). Among the PAD patients, 15 had coexisting type 2 diabetes, accounting for ca. 33% of the subjects, whereas 67%, i.e. 31 individuals, had no history of diabetes. Average glycated haemoglobin (HbA1c) level in patients with PAD and diabetes was (8.0±1.0)%. Average ankle brachial index in all subjects was 0.48±0.25 and mean intermittent claudication (IC) distance was (91.9±88.7) m. These patients also suffered from coexisting arterial hypertension (89.1%) or ischemic heart disease (43.5%), and were smokers in the vast majority (93.5%), with mean body mass index of (26.5±4.2) kg/m2.
Table 1

Characteristics of the PAD patients in the study group

Parameter

Value

Age (year)

63.0±8.5

Sex (F/M)

18 (39%)/28 (61%)

DM2+

15 (33%)

HbA1c in DM2+ subgroup

(8.0±1.0)%

DM2−

31 (67%)

Ankle brachial index

0.48±0.25

IC distance (m)

91.9±88.7

Fontaine IIa

5 (11%)

Fontaine IIb

30 (65%)

Fontaine III

3 (6.5%)

Fontaine IV

8 (17.4%)

Hypertension

41 (89.1%)

Coronary artery disease

20 (43.5%)

Smoking

43 (93.5%)

Body mass index (kg/m2)

26.5±4.2

Waist-to-hip ratio

1.0±0.1

Data are expressed as mean±standard deviation (SD) or number (percentage), with a total of 46 patients. PAD: peripheral arterial disease; DM2: type 2 diabetes; IC: intermittent claudication

Fig. 1 shows the number of patients in the PAD-DM2+ subgroup and the PAD-DM2− subgroup depending on the degree of ischemia according to Fontaine classification. The PAD-DM2+ subgroup (n =15) included 1 patient classified in IIa, 10 in IIb, and 4 in IV based on Fontaine classification. In the PAD-DM2− subgroup (n =31), there were 4 patients in IIa, 20 in IIb, 2 in III, and 5 in stage IV of disease severity.
Fig. 1

Number of patients in the subgroup with PAD and type 2 diabetes (DM2+, n =15) and the non-diabetic subgroup with PAD (DM2-, n =31) according to Fontaine classification

Table 2 displays mean concentrations of analyzed factors in the PAD-DM2+ and PAD-DM2− subgroups and in the control group. Both subgroups revealed significantly higher VEGF-A levels and lower concentrations of sVEGFR-2 than the healthy subjects.
Table 2

Plasma levels of VEGF-A, sVEGFR-1, and sVEGFR-2 in the study group (PAD) and the control group

Group

VEGF-A (pg/ml)

sVEGFR-1 (pg/ml)

sVEGFR-2 (pg/ml)

PAD (n=46)

   

  (a) DM2+ (n=15)

75.2±53.8

148.3±41.5

10987.6±3534.2

  (b) DM2− (n=31)

88.8±84.8

127.5±33.6

8753.9±2277.3

  (c) Control (n=30)

18.0±7.7

140.5±62.3

14481.5±3669.9

P-value

(a) vs. (c): 0.000 007

(a) vs. (c): 0.96

(a) vs. (c): 0.02

 

(b) vs. (c): 0.000 000 1

(b) vs. (c): 0.8

(b) vs. (c): 0.000 01

 

(a) vs. (b): 1

(a) vs. (b): 0.5

(a) vs. (b): 0.16

Date are expressed as mean±standard deviation (SD). The difference is considered to be significant at P-value of <0.05 (in bold)

Fig. 2 compares mean VEGF-A values of the PAD-DM2+ and PAD-DM2− subgroups and the control group. In the plasma of patients with PAD-DM2+, we found 4 times higher mean concentrations of VEGF-A when compared with healthy individuals (P =0.000 007). Notably, the subgroup PAD-DM2− demonstrates nearly 5 times higher VEGF-A levels against the control group (P =0.000 000 1). The differences between the PAD-DM2+ and PAD-DM2− subgroups were not statistically significant.
Fig. 2

VEGF-A levels in the subgroup of patients with PAD and type 2 diabetes (PAD-DM2+, n =15), the non-diabetic subgroup with PAD (PAD-DM2−, n =31), and the control group (n =30)

(a) vs. (c): P =0.000 007; (b) vs. (c): P =0.000 000 1; (a) vs. (b): P =1

Fig. 3 shows mean concentrations of sVEGFR-1 in the plasma of PAD-DM2+ patients, PAD-DM2 patients, and the control group, respectively. In the subgroup of PAD-DM2+, we observed slightly higher, and in the subgroup of PAD-DM2− moderately lower, levels of sVEGFR-1 in comparison with the control group. The comparative analysis of sVEGFR-1 levels between the subgroups of PAD-DM2+ and PAD-DM2− against the control group did not show any statistically significant differences.
Fig. 3

sVEGFR-2 levels in the subgroup of patients with PAD and type 2 diabetes (PAD-DM2+, n =15), the non-diabetic subgroup with PAD (PAD-DM2−, n =31), and the control group (n =30)

(a) vs. (c): P =0.02; (b) vs. (c): P =0.000 01; (a) vs. (b): P =0.16

Fig. 4 displays sVEGFR-2 levels in the subgroups of patients with PAD and in healthy subjects. In the PAD-DM2+ subgroup, we found significantly lower mean concentration of sVEGFR-2 by ca. 25% in comparison with the control group (P =0.02). The subgroup of PAD-DM2− demonstrated significantly lower sVEGFR-2 level by ca. 40% when compared with the control group (P =0.000 01). Differences in sVEGFR-2 concentrations between the two subgroups of PAD-DM2+ and PAD-DM2− were not statistically significant, yet a tendency was noted towards higher concentrations of sVEGFR-2 in PAD-DM2+ patients compared with PAD-DM2− patients.
Fig. 4

sVEGFR-2 levels in the subgroup of patients with PAD and type 2 diabetes (PAD-DM2+, n =15), the non-diabetic subgroup with PAD (PAD-DM2−, n =31), and the control group (n =30)1

(a) vs. (c): P =0.02; (b) vs. (c): P =0.000 01; (a) vs. (b): P =0.16

Obtained levels of the above mentioned factors in uniform units (pg/ml) ensured the calculation of sVEGFR-1 to VEGF-A ratio as well as sVEGFR-2 to VEGF-A ratio. Table 3 shows the figures of sVEGFR-1/VEGF-A ratio and sVEGFR-2/VEGF-A ratio in the subgroups of PAD patients and in healthy subjects. Mean sVEGFR-1/VEGF-A ratio value in the subgroup PAD-DM2+ was 71% lower than that in the control group (P =0.0002). In the subgroup of PAD-DM2−, the figure was 73% lower than that in healthy individuals (P =0.000 01). Differences between the subgroups of PAD-DM2+ and PAD-DM2− were not statistically significant. The sVEGFR-2/VEGF-A ratio was 75% lower in subgroup of PAD-DM2+ than that in the control group (P =0.000 006), whereas in the subgroup of PAD-DM2−, the figure was 82% lower than that in healthy individuals (P =0.000 001). In PAD-DM2+ patients, this parameter was insignificantly higher than that in the PAD-DM2− subgroup.
Table 3

sVEGFR-1/VEGF-A ratio and sVEGFR-2/VEGF-A ratio in the study group (PAD) and control group

Group

sVEGFR-1/VEGF-A

sVEGFR-2/VEGF-A

PAD (n=46)

  

  (a) DM2+ (n=15)

2.9±2.1

242.5±266.9

  (b) DM2− (n=31)

2.7±3.1

174.7±151.4

  (c) Control (n=30)

10.0±7.5

965.3±523.8

P-value

(a) vs. (c): 0.0002

(a) vs. (c): 0.000 006

 

(b) vs. (c): 0.000 01

(b) vs. (c): 0.000 001

 

(a) vs. (b): 1

(a) vs. (b): 1

Date are expressed as mean±standard deviation (SD). The difference is considered to be significant at P-value of <0.05 (in bold)

4 Discussion

The study revealed that PAD-DM2+ and PAD-DM2− patients had significantly higher concentrations of VEGF-A than healthy individuals (4- and 5-fold, respectively). The subgroup of PAD-DM2+ revealed slightly higher sVEGFR-1 levels and the subgroup of PAD-DM2− revealed slightly lower sVEGFR-1 levels when compared with the control group. In both subgroups, there were significantly lower sVEGFR-2 concentrations than those of healthy subjects.

The literature does not provide a uniform interpretation of angiogenesis parameters obtained in in vitro testing, animal models, or diabetic blood testing.

In the study conducted by Hochberg et al. (2001), blood samples were collected from PAD patients with coexisting diabetes or without diabetes, considering the classification based on critical limb ischemia (CLI) or chronic ischemia. In non-diabetic patients, no significant difference was observed in hypoxia-induced expression of VEGF, produced by monocytes, depending on the existence of CLI. Notably, patients with diabetes and coexisting CLI demonstrated significantly higher levels of VEGF produced by monocytes when compared with patients with diabetes but without CLI. The role of monocytes in angiogenesis and stimulation of their chemotaxis with VEGF in diabetic patients was also emphasized by Waltenberger et al. (2000) who determined VEGF levels in diabetic patients and healthy subjects, achieving significantly higher plasma concentrations of VEGF in patients with diabetes.

An animal model conducted by Hazarika et al. (2007) provided interesting results. A mouse with induced diabetes diet and artificially (surgically) induced lower limb ischemia had significantly higher baseline VEGF levels (before ischemia) in comparison with the control group. Then, after inducing ischemia, on the third day an insignificant VEGF increase was observed in mice with diabetes in comparison with non-diabetic mice, whereas on the tenth day an insignificant decrease in VEGF levels was noted in mice with diabetes as opposed to the non-diabetic group with ischemia. The authors claimed that this could suggest certain “depleting” compensation mechanisms— comprising the essence of angiogenesis and collateral circulation. Our study revealed slightly lower VEGF concentrations in the subgroup of PAD-DM2+, compared with PAD-DM2− patients with chronic ischemia.

Attempts to explain complex angiogenesis processes in patients with diabetes were made by Chung et al. (2006) who performed internal mammary artery biopsy in 32 diabetic patients and 32 non-diabetic patients and, apart from the levels of VEGF, they determined angiostatin and metalloproteinases (MMP-2 and MMP-9). They found significantly lower VEGF expression in patients suffering from type 2 diabetes in comparison with the control group and emphasized the involvement of other above-mentioned factors in angiogenesis. Libra et al. (2006) analyzed the impact of genetic polymorphism on interleukin-6 (IL-6) which affects VEGF secretion. Specific IL-6 genotypes were identified in individuals with type 2 diabetes coexistent with PAD, which induced higher VEGF levels in diabetic patients with PAD.

Blann et al. (2002) assessed the expression of angiogenic factors in plasma of patients with type 2 diabetes, and revealed significantly higher VEGF concentrations in diabetic patients comparable with healthy subjects. Similarly higher VEGF concentrations in plasma of patients with PAD irrespective of diabetes (non-)coexistence were reported by Botti et al. (2012). Yet Zakareia (2012) observed much higher plasma VEGF levels in subjects with diabetes and PAD. In the study on patients with the DFS, Drela et al. (2014) reported significantly higher VEGF-A concentrations in patients with type 2 diabetes. Ruszkowska-Ciastek et al. (2014) observed non-significant differences in the serum concentrations of VEGF-A between the patients with well-controlled diabetes and the control group. Orrico et al. (2010) carried out a study involving 33 subjects with CLI, identifying a subgroup of 22 type 2 diabetic patients. Notably, they revealed higher VEGF levels in plasma of all patients with CLI compared with the control group, whereas no differences were observed between the subgroups of diabetic and non-diabetic patients. The study of 70 subjects suffering from diabetes coexistent with PAD carried out by Bleda et al. (2012) revealed higher plasma VEGF levels in individuals with CLI compared with patients with chronic ischemia.

Based on the studies outlined above, patients with PAD and coexisting type 2 diabetes have higher VEGF-A levels. Undeniably, the crucial factor affecting high concentrations of VEGF-A, regardless of the presence of type 2 diabetes in patients with PAD, is hypoxia caused by stenosis or obstruction of “large and medium” caliber arteries due to the presence of atherosclerotic plaques. The achieved findings suggest that the coexistence of type 2 diabetes may inhibit angiogenesis, which is reflected by lower VEGF-A levels than that in arterial disease alone, as demonstrated in this study. This may be connected with the currently adopted view that hyperglycemia has a considerable impact on functional and structural damage to vascular endothelial cells—the so called microangiopathic complications (Duh and Aiello, 1999). Moreover, it is postulated that the stock of EPCs, responsible for vessel repair mechanisms, runs down much more in patients with diabetes than in patients with arterial disease, while hyperglycemia, observed in patients with diabetes, can damage chemotaxis and migration of EPCs (Fadini et al., 2005; Orrico et al., 2010). Therefore, not only VEGF therapy but also administration of mononuclear cells is included in the treatment of CLI (Skóra et al., 2013). To evaluate the role of VEGF in the angiogenic process in PAD and diabetes, the contribution of such factors as angiopoietin-1 and angiopoietin-2 should also be taken into consideration (Lim et al., 2005). sVEGFR-1 is a recognized angiogenesis inhibitor. Soluble receptor (sVEGFR-1) and stationary VEGFR-1 anchored in endothelial cell membrane comprise a significant mechanism that inhibits VEGF circulating in the blood, and are referred to as decoy receptors (Meyer et al., 2006). Their role is to catch VEGF-A ensuring a significant decrease in its bioavailability to VEGFR-2 on the surface of endothelial cells, reduce stimulation of cell signaling pathways, and therefore decrease biological function of VEGF. Our study showed slightly higher mean concentrations of sVEGFR-1 in the subgroup of patients with PAD and coexisting type 2 diabetes in comparison with the control group. Hazarika et al. (2007) conducted a study on an animal model to test sVEGFR-1 concentrations. Initial marking of sVEGFR-1 (before inducing ischemia) in the blood of mice with diabetes revealed significantly lower values comparable with non-diabetic mice. Then, on the third and tenth days following ischemia induction, sVEGFR-1 levels grew in the diabetic group and were higher than those in the non-diabetic subgroup.

Although this study did not reveal significantly different sVEGFR-1 levels in both subgroups of patients suffering from PAD, the inhibition capacity of VEGF decreased noticably in comparison with healthy individuals, from a 10-fold sVEGFR-1 predominance to ca. 3-fold in sick subjects. In this study we noted lower mean values of sVEGFR-2 in both subgroups of PAD-DM2+ and PAD-DM2− compared with the control group. Moreover, slightly higher sVEGFR-2 levels were observed in the subgroup with PAD and diabetes comparable with the non-diabetic subgroup with PAD. Similar results (versus the control group), i.e. decreased mean concentrations of sVEGFR-2 in patients with DFS, were achieved by Drela et al. (2014). No significant differences in the serum concentration of (s)VEGFR1 or (s)VEGFR-2 were observed by Ruszkowska-Ciastek et al. (2014) between the subjects with diabetes and the control group.

As shown in Table 3, the figures of sVEGFR-1/VEGF-A ratio in both subgroups of patients with PAD and in the control group indicated that the inhibition capacity expressed by the value of the above ratio is lower in patients with PAD than in healthy individuals despite significantly higher VEGF-A levels, which indicates the role of receptor type 1 in this process. Available studies do not provide information to assess the concentrations of formed sVEGFR-1–VEGF-A complexes. However, it should be noted that relatively low angiogenic inhibition capacity in both subgroups with PAD vs. the control group is the result of sVEGFR-1 being used up in the binding processes of VEGF-A when the levels of VEGF-A were high. Therefore, sVEGFR-1/VEGF-A ratio well illustrates decreased bioavailability of VEGF-A in patients with lower limb ischemia.

There are no studies on the evaluation of mutual proportions of VEGF-A and sVEGFR-1 levels or VEGF-A and sVEGFR-2 levels in patients suffering from PAD and coexisting diabetes. VEGF-A/sVEGFR-1 index was assessed in patients with pancreatic cancer and acute myeloid leukemia with regard to the severity of the disease and prognosis (Aref et al., 2005; Chang et al., 2008). This parameter could be used in practice to assess the degree of severity of PAD, which is the subject of the current research (Rość et al., 2014). The studies by Toi et al. (2002) involving patients with breast cancer and the studies by Yamaguchi et al. (2007) concerning individuals suffering from colorectal carcinoma, were focused on analyzing the sVEGFR-1/VEGF ratio in terms of clinical parameters and prognosis of cancer. This study showed stronger angiogenic inhibition capacity demonstrated by the ratio of concentrations (sVEGFR-1/VEGF-A and sVEGFR-2/VEGF-A) in the control group in comparison with PAD subgroups. Yet the subgroup of patients with PAD and coexisting diabetes revealed a tendency towards higher anti-angiogenic potential comparable with non-diabetic patients with PAD, better illustrated by sVEGFR-2/VEGF-A ratio than by sVEGFR-1/VEGF-A ratio. This may have clinical significance and suggest increased but impaired angiogenesis in diabetic individuals, which may explain, for example, impaired healing in the DFS. Nevertheless, this certainly needs to be confirmed by large-scale studies of patients with PAD and diabetes. This study should be treated as preliminary because there were insufficient numbers of patients in the study groups and the subgroup of non-diabetic patients with PAD was twice the size of the subgroup of type 2 diabetic subjects with PAD.

5 Conclusions

The coexistence of type 2 diabetes and PAD is demonstrated by a tendency to lower plasma levels of proangiogenic factor (VEGF-A) and higher levels of angiogenesis inhibitors (sVEGFR-1 and sVEGFR-2) at the same time. Regardless of the coexistence of type 2 diabetes, hypoxia appears to be a crucial factor stimulating the processes of angiogenesis in PAD patients comparable with healthy individuals, whereas hyperglycemia may have a negative (inhibitive) impact on angiogenesis in lower limbs.

Compliance with ethics guidelines

Radosław WIECZÓR, Grażyna GADOMSKA, Barbara RUSZKOWSKA-CIASTEK, Katarzyna STANKOWSKA, Jacek BUDZYŃSKI, Jacek FABISIAK, Karol SUPPAN, Grzegorz PULKOWSKI, and Danuta ROŚĆ declare that they have no conflict of interest.

All procedures followed were in accordance with the ethical standards of the responsible committee on human experimentation (institutional and national) and with the Helsinki Declaration of 1975, as revised in 2008 (5). Informed consent was obtained from all patients for being included in the study. Additional informed consent was obtained from all patients for whom identifying information is included in this article.

References

  1. Aref, S., El Sherbiny, M., Goda, T., et al., 2005. Soluble VEGF/sFLt1 ratio is an independent predictor of AML patient outcome. Hematology, 10 (2): 131–134. [doi:10.1080/10245330500065797]CrossRefPubMedGoogle Scholar
  2. Bao, P., Kodra, A., Tomic-Canic, M., et al., 2009. The role of vascular endothelial growth factor in wound healing. J. Surg. Res., 153 (2): 347–358. [doi:10.1016/j.jss.2008.04.023]CrossRefPubMedGoogle Scholar
  3. Blann, A.D., Belgore, F.M., McCollum, C.N., et al., 2002. Vascular endothelial growth factor and its receptor, Flt-1, in the plasma of patients with coronary or peripheral atherosclerosis, or Type IIdiabetes. Clin. Sci., 102: 187-194. [doi:10.1042/cs1020187]Google Scholar
  4. Bleda, S., de Haro, J., Varela, C., et al., 2012. Impact of VEGF polymorphisms on the severity of peripheral artery disease in diabetic patients. Growth Factors, 30 (5): 277–282. [doi:10.3109/08977194.2012.703664]CrossRefPubMedGoogle Scholar
  5. Botti, C., Maione, C., Dogliotti, G., et al., 2012. Circulating cytokines present in the serum of peripheral arterial disease patients induce endothelial dysfunction. J. Biol. Regul. Homeost. Agents, 26 (1): 67–79.PubMedGoogle Scholar
  6. Chang, Y.T., Chang, M.C., Wei, S.C., et al., 2008. Serum vascular endothelial growth factor/soluble vascular endothelial growth factor receptor 1 ratio is an independent prognostic marker in pancreatic cancer. Pancreas, 37(2): 145–150. [doi:10.1097/MPA.0b013e318164548a]CrossRefPubMedGoogle Scholar
  7. Chung, A.W., Hsiang, Y.N., Matzke, L.A., et al., 2006. Reduced expression of vascular endothelial growth factor paralleded with the increased angiostatin expression resulting from the upregulated activities matrix metalloproteinase-2 and -9 in human type 2 diabetic arterial vasculature. Circ. Res., 99 (2): 140–148. [doi:10.1161/01. RES.0000232352.90786.fa]CrossRefPubMedGoogle Scholar
  8. Drela, E., Kulwas, A., Jundzill, A., et al., 2014. VEGF-A and PDGF-BB—angiogenic factors and the stage of diabetic foot syndrome advancement. Endokrynol. Pol., 65(4): 306–312. [doi:10.5603/EP.2014.0042]CrossRefPubMedGoogle Scholar
  9. Duh, E., Aiello, L.P., 1999. Vascular endothelial growth factor and diabetes: the agonist versus antagonist paradox. Diabetes, 48 (10): 1899–1906. [doi:10.2337/diabetes.48.10. 1899]CrossRefPubMedGoogle Scholar
  10. Fadini, G.P., Miorin, M., Facco, M., et al., 2005. Circulating endothelial progenitor cells are reduced in peripheral vascular complications of type 2 diabetes mellitus. J. Am. Coll. Cardiol., 45 (9): 1449–1457. [doi:10.1016/j.jacc. 2004.11.067]CrossRefPubMedGoogle Scholar
  11. Hazarika, S., Dokun, A.O., Li, Y., et al., 2007. Impaired angiogenesis after hindlimb ischemia in type 2 diabetes mellitus: differential regulation of vascular endothelial growth factor receptor 1 and soluble vascular endothelial growth factor receptor 1. Circ. Res., 101 (9): 948–956. [doi:10.1161/CIRCRESAHA.107.160630]CrossRefPubMedGoogle Scholar
  12. Hochberg, I., Hoffman, A., Levy, A.P., 2001. Regulation of VEGF in diabetic patients with critical limb ischemia. Ann. Vasc. Surg., 15 (3): 388–392. [doi:10.1007/s1001600 10089]CrossRefPubMedGoogle Scholar
  13. Jude, E.B., Eleftheriadou, I., Tentolouris, N., 2010. Peripheral arterial disease in diabetes—a review. Diabetes Med., 27 (1): 4–14. [doi:10.1111/j.1464-5491.2009.02866.x]CrossRefGoogle Scholar
  14. Kusumanto, Y.H., van Weel, V., Mulder, N.H., et al., 2006. Treatment with intramuscular vascular endothelial growth factor gene compared with placebo for patients with diabetes mellitus and critical limb ischemia: a double-blind randomized trial. Hum. Gene Ther., 17(6): 683-691. [doi:10.1089/hum.2006.17.683]Google Scholar
  15. Libra, M., Signorelli, S.S., Bevelacqua, Y., et al., 2006. Analysis of G(-174)C IL-6 polymorphism and plasma concentrations of inflammatory markers in patients with type 2 diabetes and peripheral arterial disease. J. Clin. Pathol., 59 (2): 211–215. [doi:10.1136/jcp.2004.025452]CrossRefPubMedPubMedCentralGoogle Scholar
  16. Lim, H.S., Lip, G.Y., Blann, A.D., 2005. Angiopoietin-1 and angiopoietin-2 in diabetes mellitus: relationship to VEGF, glycaemic control, endothelial damage/dysfunction and atherosclerosis. Atherosclerosis, 180 (1): 113–118. [doi:10.1016/j.atherosclerosis.2004.11.004]CrossRefPubMedGoogle Scholar
  17. Liu, W., Zhang, X., Song, C., et al., 2014. Expression and characterization of a soluble VEGF receptor 2 protein. Cell Biosci., 4(1):14. [doi:10.1186/2045-3701-4-14]CrossRefPubMedPubMedCentralGoogle Scholar
  18. Meyer, R.D., Mohammadi, M., Rahimi, N., 2006. A single amino acid substitution in the activation loop defines the decoy characteristic of VEGFR-1/FLT-1. J. Biol. Biochem., 281 (2): 867–875. [doi:10.1074/jbc.m506454200]Google Scholar
  19. Norgren, L., Hiatt, W.R., Dormandy, J.A., et al., 2007. Intersociety consensus for the management of peripheral arterial disease (TASC II). Eur. J. Vasc. Endovasc. Surg., 33(Suppl. 1):S1–S75. [doi:10.1016/j.ejvs.2006.09.024]CrossRefPubMedGoogle Scholar
  20. Orrico, C., Pasquinelli, G., Foroni, L., et al., 2010. Dysfunctional vasa vasorum in diabetic peripheral artery obstructive disease with critical lower limb ischaemia. Eur. J. Vasc. Endovasc. Surg., 40 (3): 365–374. [doi:10.1016/j. ejvs.2010.04.011]CrossRefPubMedGoogle Scholar
  21. Rosc, D., Wieczór, R., Stankowska, K., et al., 2014. Plasma VEGF-A/SVEGFR-1 ratio as a potential ischemic marker in patients with symptomatic peripheral arterial disease— preliminary report. Thromb. Res., 133(Suppl. 3):95. [doi: 10.1016/S0049-3848(14)50305-2]CrossRefGoogle Scholar
  22. Ruszkowska-Ciastek, B., Sokup, A., Socha, M.W., et al., 2014. A preliminary evaluation of VEGF-A, VEGFR1 and VEGFR2 in patients with well-controlled type 2 diabetes mellitus. J. Zhejiang Univ.-Sci. B (Biomed. & Biotechnol.), 15 (6): 575–581. [doi:10.1631/jzus.B1400024]CrossRefGoogle Scholar
  23. Skóra, J., Barc, P., Pupka, A., et al., 2013. Transplantation of autologous bone marrow mononuclear cells with VEGF gene improves diabetic critical limb ischaemia. Endokrynol. Pol., 64 (2): 129–138.PubMedGoogle Scholar
  24. Toi, M., Bando, H., Ogawa, T., et al., 2002. Significance of vascular endothelial growth factor (VEGF)/soluble VEGF receptor-1 relationship in breast cancer. Int. J. Cancer, 98 (1): 14–18. [doi:10.1002/ijc.10121]CrossRefPubMedGoogle Scholar
  25. Waltenberger, J., Lange, J., Kranz, A., 2000. Vascular endothelial growth factor-A induced chemotaxis of monocytes is attenuated in patients with diabetes mellitus: a potential predictor for the individual capacity to develop collaterals. Circulation, 102 (2): 185–190. [doi:10.1161/01.CIR.102. 2.185]CrossRefPubMedGoogle Scholar
  26. Yamaguchi, T., Bando, H., Mori, T., et al., 2007. Overexpression of soluble vascular endothelial growth factor receptor 1 in colorectal cancer: association with progression and prognosis. Cancer Sci., 98 (3): 405–410. [doi:10.1111/j.1349-7006.2007.00402.x]CrossRefPubMedGoogle Scholar
  27. Zakareia, F.A., 2012. Correlation of peripheral arterial blood flow with plasma chemerin and VEGF in diabetic peripheral vascular disease. Biomark. Med., 6 (1): 81–87. [doi:10.2217/bmm.11.85]CrossRefPubMedGoogle Scholar

Copyright information

© Zhejiang University and Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  • Radosław Wieczór
    • 1
    • 2
    Email author
  • Grażyna Gadomska
    • 1
  • Barbara Ruszkowska-Ciastek
    • 1
  • Katarzyna Stankowska
    • 1
  • Jacek Budzyński
    • 2
    • 3
  • Jacek Fabisiak
    • 2
  • Karol Suppan
    • 2
  • Grzegorz Pulkowski
    • 2
  • Danuta Rość
    • 1
  1. 1.Department of Pathophysiology, Faculty of PharmacyNicolaus Copernicus University in Toruń, Ludwik Rydygier Collegium Medicum in BydgoszczBydgoszczPoland
  2. 2.Clinic of Vascular and Internal MedicineDr. Jan Biziel University HospitalBydgoszczPoland
  3. 3.Department of Vascular and Internal Medicine, Faculty of Health Sciences, Nicolaus Copernicus University in ToruńLudwik Rydygier Collegium Medicum in BydgoszczBydgoszczPoland

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