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. 2015 Dec 28;12(1):175–190. doi: 10.1007/s11302-015-9491-2

Table 2.

Kinetic parameters for L-arginine transport in HUVECs from normal or GDM pregnancies

Saturable transport Overall transport
V max (pmol/μg protein/ min) K m (μM) V max/K m (pmol/μg protein/min/(μM)) K D (pmol/μg protein/min/(μM)) v i (pmol/μg protein/ 0.5 s)
Normal pregnancies
 Without insulin
  Control 1.2 ± 0.2 89 ± 20 0.0135 ± 0.0027 0.0011 ± 0.0004 0.006 ± 0.0001
  A1AR ag 1.2 ± 0.1 103 ± 21 0.0117 ± 0.0016 0.0012 ± 0.0005 0.006 ± 0.0001
  A1AR antag 1.0 ± 0.2 77 ± 40 0.0130 ± 0.0047 0.0011 ± 0.0004 0.005 ± 0.0002
  A1AR ag/antag 1.1 ± 0.3 79 ± 26 0.0139 ± 0.0042 0.0012 ± 0.0003 0.006 ± 0.0002
  A2AAR ag 1.1 ± 0.2 108 ± 15 0.0102 ± 0.0016 0.0011 ± 0.0003 0.005 ± 0.0001
  A2AAR antag 1.0 ± 0.1 99 ± 33 0.0101 ± 0.0022 0.0010 ± 0.0003 0.007 ± 0.0001
  A2AAR ag/antag 1.3 ± 0.4 87 ± 24 0.0149 ± 0.0044 0.0013 ± 0.0007 0.008 ± 0.0003
  A2BAR ag 1.5 ± 0.3 98 ± 21 0.0153 ± 0.0031 0.0012 ± 0.0004 0.008 ± 0.0002
  A2BAR antag 1.2 ± 0.1 91 ± 16 0.0132 ± 0.0017 0.0011 ± 0.0004 0.006 ± 0.0001
  A2BAR ag/antag 1.1 ± 0.3 87 ± 21 0.0126 ± 0.0032 0.0012 ± 0.0006 0.006 ± 0.0002
  A3AR ag 0.9 ± 0.2 62 ± 23 0.0145 ± 0.0043 0.0012 ± 0.0005 0.006 ± 0.0002
  A3AR antag 1.3 ± 0.1 97 ± 21 0.0134 ± 0.0020 0.0013 ± 0.0004 0.007 ± 0.0001
  A3AR ag/antag 1.2 ± 0.3 96 ± 31 0.0125 ± 0.0036 0.0012 ± 0.0005 0.006 ± 0.0002
 With insulin
  Control 2.3 ± 0.1* 93 ± 22 0.0247 ± 0.0035* 0.0012 ± 0.0004 0.012 ± 0.0002*
  A1AR ag 2.7 ± 0.1* 104 ± 21 0.0260 ± 0.0031* 0.0012 ± 0.0004 0.013 ± 0.0003*
  A1AR antag 2.2 ± 0.1* 88 ± 20 0.0250 ± 0.0035* 0.0011 ± 0.0005 0.011 ± 0.0003*
  A1AR ag/antag 2.2 ± 0.2* 81 ± 16 0.0272 ± 0.0039* 0.0012 ± 0.0006 0.012 ± 0.0003*
  A2AAR ag 2.9 ± 0.2* 137 ± 20 0.0212 ± 0.0023* 0.0010 ± 0.0004 0.011 ± 0.0002*
  A2AAR antag 0.9 ± 0.4† 95 ± 29 0.0095 ± 0.0036† 0.0011 ± 0.0004 0.004 ± 0.0002†
  A2AAR ag/antag 1.1 ± 0.2† 82 ± 27 0.0134 ± 0.0034† 0.0013 ± 0.0005 0.007 ± 0.0002†
  A2BAR ag 1.7 ± 0.1* 95 ± 21 0.0179 ± 0.0025* 0.0010 ± 0.0005 0.011 ± 0.0002*
  A2BAR antag 2.3 ± 0.2* 110 ± 29 0.0209 ± 0.0031* 0.0013 ± 0.0003 0.012 ± 0.0002*
  A2BAR ag/antag 1.9 ± 0.3* 89 ± 21 0.0213 ± 0.0043* 0.0013 ± 0.0005 0.011 ± 0.0003*
  A3AR ag 2.5 ± 0.1* 84 ± 12 0.0298 ± 0.0027* 0.0012 ± 0.0005 0.014 ± 0.0003*
  A3AR antag 2.2 ± 0.1* 88 ± 18 0.0250 ± 0.0033* 0.0011 ± 0.0003 0.011 ± 0.0002*
  A3AR ag/antag 2.4 ± 0.3* 99 ± 18 0.0242 ± 0.0038* 0.0013 ± 0.0006 0.013 ± 0.0003*
GDM pregnancies
 Without insulin
  Control 4.3 ± 0.3 120 ± 31 0.0358 ± 0.0059 0.0013 ± 0.0003 0.021 ± 0.004
  A1AR ag 4.3 ± 0.3 143 ± 33 0.0301 ± 0.0045 0.0014 ± 0.0004 0.021 ± 0.004
  A1AR antag 4.2 ± 0.3 125 ± 25 0.0336 ± 0.0045 0.0012 ± 0.0002 0.019 ± 0.003
  A1AR ag/antag 4.1 ± 0.5 119 ± 34 0.0345 ± 0.0071 0.0013 ± 0.0003 0.020 ± 0.004
  A2AAR ag 4.0 ± 0.3 150 ± 35 0.0267 ± 0.0041 0.0012 ± 0.0001 0.016 ± 0.002
  A2AAR antag 2.1 ± 0.5† 135 ± 42 0.0156 ± 0.0043† 0.0012 ± 0.0001 0.009 ± 0.002†
  A2AAR ag/antag 2.2 ± 0.4† 127 ± 26 0.0173 ± 0.0034† 0.0011 ± 0.0003 0.010 ± 0.002†
  A2BAR ag 4.3 ± 0.3 134 ± 33 0.0321 ± 0.0051 0.0012 ± 0.0001 0.018 ± 0.002
  A2BAR antag 4.1 ± 0.3 120 ± 34 0.0342 ± 0.0060 0.0012 ± 0.0002 0.019 ± 0.003
  A2BAR ag/antag 3.9 ± 0.6 129 ± 37 0.0302 ± 0.0067 0.0012 ± 0.0004 0.017 ± 0.004
  A3AR ag 4.1 ± 0.3 142 ± 53 0.0289 ± 0.0064 0.0012 ± 0.0001 0.017 ± 0.003
  A3AR antag 4.2 ± 0.3 135 ± 43 0.0311 ± 0.0061 0.0012 ± 0.0001 0.018 ± 0.003
  A3AR ag/antag 4.0 ± 0.5 121 ± 31 0.0331 ± 0.0064 0.0013 ± 0.0003 0.020 ± 0.004
 With insulin
  Control 2.2 ± 0.5* 172 ± 40 0.0128 ± 0.0029* 0.0013 ± 0.0003 0.009 ± 0.002*
  A1AR ag 2.2 ± 0.4* 142 ± 35 0.0155 ± 0.0033* 0.0014 ± 0.0040 0.011 ± 0.012*
  A1AR antag 4.4 ± 0.3† 162 ± 46 0.0272 ± 0.0048† 0.0012 ± 0.0002 0.017 ± 0.003†
  A1AR ag/antag 4.1 ± 0.4† 149 ± 35 0.0275 ± 0.0047† 0.0013 ± 0.0003 0.018 ± 0.003†
  A2AAR ag 1.9 ± 0.3* 134 ± 31 0.0142 ± 0.0028* 0.0012 ± 0.0001 0.008 ± 0.001*
  A2AAR antag 1.9 ± 0.4* 163 ± 95 0.0117 ± 0.0046* 0.0012 ± 0.0001 0.007 ± 0.002*
  A2AAR ag/antag 2.2 ± 0.4* 135 ± 25 0.0163 ± 0.0030* 0.0013 ± 0.0003 0.010 ± 0.002*
  A2BAR ag 2.2 ± 0.2* 142 ± 37 0.0155 ± 0.0027* 0.0012 ± 0.0001 0.009 ± 0.001*
  A2BAR antag 2.0 ± 0.3* 173 ± 72 0.0116 ± 0.0033* 0.0012 ± 0.0001 0.007 ± 0.002*
  A2BAR ag/antag 2.2 ± 0.3* 165 ± 25 0.0133 ± 0.0019* 0.0014 ± 0.0005 0.010 ± 0.002*
  A3AR ag 2.3 ± 0.2* 162 ± 34 0.0142 ± 0.0021* 0.0012 ± 0.0001 0.009 ± 0.001*
  A3AR antag 2.3 ± 0.3* 163 ± 53 0.0141 ± 0.0032* 0.0012 ± 0.0001 0.009 ± 0.002*
  A3AR ag/antag 2.2 ± 0.2* 152 ± 25 0.0145 ± 0.0018* 0.0011 ± 0.0005 0.008 ± 0.002*

Data are presented as mean ± SEM (n = 38). L-Arginine transport (0–1000 μM, 1 min, 37 °C) was measured in HUVECs from normal (Normal) or gestational diabetes mellitus (GDM) pregnancies exposed (8 h) to culture medium without (Without insulin) or with (With insulin) insulin (1 nM). Transport assays were done in the absence (Control) or presence of the A1AR agonist CPA (30 nM, A1AR ag), antagonist DPCPX (30 nM, A1AR antag), or both (A1AR ag/antag), the A2AAR agonist CGS-21680 (30 nM, A2AAR ag), antagonist ZM-241385 (10 nM, A2AAR antag), or both (A2AAR ag/antag), the A2BAR agonist BAY60-6583 (100 nM, A2BAR agonist), antagonist MRS-1754 (30 nM, A2BAR antag), or both (A2BAR ag/antag), the A3AR agonist 2-Cl-IB-MECA (30 nM, A3AR ag), antagonist MRS-1523 (30 nM, A3AR antag) or both (A3AR ag/antag) (see “Materials and methods”). Maximal velocity (V max) and apparent Michaelis-Menten constant (K m) of saturable transport were calculated assuming a single Michaelis-Menten hyperbola. V max/K m represents maximal L-arginine transport capacity. The lineal phase of overall transport of L-arginine (K D) was obtained from transport data fitted to a Michaelis-Menten equation increased in a lineal component. Initial velocity (v i) was calculated for 0.5 s with 100 μM L-arginine transport

*P < 0.05 versus corresponding values in Without insulin; †P < 0.05 versus corresponding Control values