Biomarkers for diabetes, obesity, and/or hypertension

A method of determining the level of angiotensinogen or apolipoprotein C1 in a test plasma sample relative to a control plasma sample, the method comprising:

• obtaining a test plasma sample from a test subject and a control plasma sample from a control subject;
• immuno-depleting albumin, IgG, IgA, transferrin, antitrypsin, and haptoglobin (i) from the test plasma sample, resulting in a depleted test plasma sample, and (ii) from the control plasma sample, resulting in a depleted control plasma sample;
• separating (i) the depleted test plasma sample into a first fraction comprising glycosylated polypeptides and a second fraction comprising non-glycosylated polypeptides, and (ii) the depleted control plasma sample into a third fraction comprising glycosylated polypeptides and a fourth fraction comprising non-glycosylated polypeptides;
• digesting with trypsin (i) the non-glycosylated polypeptides in the second fraction, resulting in a test tryptic digest, and (ii) the non-glycosylated polypeptides in the fourth fraction, resulting in a control tryptic digest;
• subjecting to liquid chromatography-mass spectrometry (LC/MS) (i) the test tryptic digest, resulting in a test polypeptide profile, and (ii) the control tryptic digest, resulting in a control polypeptide profile;
• and
• comparing the level of angiotensinogen or apolipoprotein C1 in the test polypeptide profile to the level of angiotensinogen or apolipoprotein C1 in the control polypeptide profile.

A method of identifying a biomarker for diabetes, obesity, and/or hypertension, the method comprising:

• obtaining a test plasma sample from a subject having or at risk of developing diabetes, obesity, and/or hypertension, and obtaining a control plasma sample from a subject not having or not at risk of developing diabetes, obesity, and/or hypertension;
• immuno-depleting albumin, IgG, IgA, transferrin, antitrypsin, and haptoglobin from the test and control samples, resulting in a depleted test sample and a depleted control sample;
• separating the depleted test sample into a first fraction comprising test glycosylated polypeptides and a second fraction comprising test non-glycosylated polypeptides, and separating the depleted control sample into a third fraction comprising control glycosylated polypeptides and a fourth fraction comprising control non-glycosylated polypeptides;
• independently digesting the test non-glycosylated polypeptides in the second fraction and the control non-glycosylated polypeptides in the fourth fraction with trypsin, resulting in a test tryptic digest and a control tryptic digest;
• independently subjecting the test tryptic digest and the control tryptic to LC/MS, resulting in a test polypeptide profile and a control polypeptide profile;
• and
• comparing the test polypeptide profile and the control polypeptide profile,
• a polypeptide present at a level in the test polypeptide profile that is different than in the control polypeptide profile being indicative of the polypeptide as a biomarker for diabetes, obesity, and/or hypertension.

A method of identifying a subject having, or at risk of developing obesity and/or comorbid diabetes and hypertension, the method comprising:

• determining the level of angiotensinogen or apolipoprotein C1 (i) in a test plasma sample obtained from the subject and (ii) in a control plasma sample of like tissue derivation from a control subject not having, or at risk of developing, diabetes, obesity, and/or hypertension wherein the step of determining the level of angiotensinogen or apolipoprotein C1 in the test plasma sample and in the control plasma sample comprises:
• immuno-depleting albumin, IgG, IgA, transferrin, antitrypsin, and haptoglobin (i) from the test plasma sample, resulting in a depleted test plasma sample, and (ii) from the control plasma sample, resulting in a depleted control plasma sample;
• separating (i) the depleted test plasma sample into a first fraction comprising glycosylated polypeptides and a second fraction comprising non-glycosylated polypeptides, and (ii) the depleted control plasma sample into a third fraction comprising glycosylated polypeptides and a fourth fraction comprising non-glycosylated polypeptides;
• digesting with trypsin (i) the non-glycosylated polypeptides in the second fraction, resulting in a test tryptic digest, and (ii) the non-glycosylated polypeptides in the fourth fraction, resulting in a control tryptic digest;
• and subjecting to liquid chromatography-mass spectrometry (LC/MS) (i) the test tryptic digest, resulting in a test polypeptide profile, and (ii) the control tryptic digest, resulting in a control polypeptide profile;
• and
• comparing the level of angiotensinogen or apolipoprotein C1 in the test sample and in the control sample, wherein an increased level of angiotensinogen or apolipoprotein C1 in the test sample relative to the control sample being indicative that the subject has, or is at risk of developing obesity and/or comorbid diabetes and hypertension.